Xu-Wei Chen

Selective extraction/isolation of hemoglobin with ionic liquid 1-butyl-3-trimethylsilylimidazolium hexafluorophosphate (BtmsimPF6)

Ionic liquid was for the first time employed for selective isolation of heme-protein species. Direct extraction of hemoglobin into ionic liquid without using any concomitant reagent or extractant was carried out. Hemoglobin at the level of 100 ng microL(-1) could readily be quantitatively extracted into ionic liquid (IL) 1-butyl-3-trimethylsilylimidazolium hexafluorophosphate (BtmsimPF(6)) in the absence of any co-existing extractants/additives at pH 7, at the same time; however, the other protein species do not interfere and remain in the aqueous phase. A back extraction efficiency of ca. 80% for 20 ng microL(-1) hemoglobin in ionic liquid phase was achieved with sodium dodecyl sulfate (SDS) solution as stripping reagent. (57)Fe Mossbauer spectra and circular dichroism (CD) spectra indicated that the penta-coordinated ferrous atom in hemoglobin provide a vacant or free coordinating position, which could be occupied by the cationic Btmsim(+) moiety. The interaction/coordination reaction between the iron atom in the heme group of hemoglobin and the cationic ionic liquid moiety furnishes the driving force for facilitating fast transfer of hemoglobin into BtmsimPF(6). The present system was applied for selective isolation of heme-protein, i.e., hemoglobin from human whole blood without any pretreatment, giving rise to satisfactory results.

Ionic liquid-polyvinyl chloride ionomer for highly selective isolation of basic proteins.

Hydrophilic ionic liquid-polyvinyl chloride (PVC) hybrids were prepared by immobilizing N-methylimidazole (N-mim) to PVC chains in toluene. The NmimCl-PVC hybrids were characterized by FT-IR, (1)H NMR, surface charge analysis and elemental analysis. The immobilization ratio, i.e., the percentage of chloride on PVC chain reacting with N-mim to form the hybrid, varies from 4.3% to 15.1% by increasing the N-mim/PVC molar ratio. The most distinct feature of the hybrid is its excellent selectivity for adsorbing basic proteins by effective suppression of the non-specific protein adsorption by pure PVC, and a higher immobilization ratio facilitates better selectivity. In Tris-HCl buffer, 100 microg mL(-1) of basic proteins, i.e., lysozyme (Lys), cytochrome c (cyt-c) and hemoglobin (Hb), were favorably adsorbed with efficiencies of 97%, 98% and 94% by the hybrid with an immobilization ratio of 15.1%, while the adsorption of acidic proteins, i.e., bovine albumin serum (BSA), transferring (Trf) and immunoglobulin G (IgG) were negligible. The retained Lys, cyt-c and Hb could be readily recovered by elution with phosphate buffer, carbonate buffer and SDS solution with efficiencies of 89%, 87% and 84%, respectively. Another feature of the hybrid is the significant improvement of the biocompatibility characterized by the maintenance of the activity of hemoglobin after adsorption and elution process. The practical usefulness of the hybrid was demonstrated by selective isolation of hemoglobin from human whole blood.

Silk fibroin as a sorbent for on-line extraction and preconcentration of copper with detection by electrothermal atomic absorption spectrometry.

Silk fibroin is a kind of polypeptide with functional amino acids in its structure. The electric charges in its molecular chains originating from the dissociation of acidic groups, i.e., hydroxyl, phenol and carboxyl, provide vast potentials for the retention of metal species of interest. In this study, the selective retention of Cu(2+) with silk fibroin at pH 6.0 was investigated and a novel on-line procedure for separation/preconcentration of Cu(2+) from complex sample matrices was thus developed by using a sequential injection system with an electrothermal atomic absorption spectrometry. A novel concept of enrichment index (EI), i.e., defined as enrichment factor (EF) obtained by consuming unity of sample volume (ml), was proposed for evaluating the enrichment efficiency of a flow-based preconcentration procedure. With a sampling volume of 900 microl, an EI of 30.3 (EF=27.3) was achieved, which was much improved as compared to that of reported procedures. A detection limit of 8.0 ng l(-1) was achieved within a linear range of 0.025-1.5 microg l(-1) along with a precision of 2.2% R.S.D. at 0.5 microg l(-1). The practical applicability of this procedure was validated by analyzing a certified reference material of riverine water (GBW08608) and a certified reference material of seawater (NASS-5) achieving satisfactory agreements between the certified and the obtained values. A spiking recovery was also performed by using a cave water sample.

A sequential injection fluorometric procedure for rapid determination of total protein in human serum

A sensitive procedure for the quantification of total protein bovine serum albumen (BSA) in human serum was presented with sequential injection sampling and fluorometric detection. A few microliters of sample and fluorescamine solutions were aspirated into the holding coil to facilitate the reaction of protein with fluorescamine by giving rise to a blue-green-fluorescent derivative. The derivative was afterwards excited by a 400nm radiation from a UV radiator, and the emitted fluorescence was monitored at the wavelength of 470nm. By loading 5.0mul of sample and 4.0mul of fluorescamine solution 0.075% (m/v), a linear calibration graph was obtained within 0.3-12.5mugml(-1), and a detection limit (3sigma) of 0.1mugml(-1) was achieved, along with a sampling frequency of 40h(-1) and a R.S.D. value of 2.1% at the 5.0mugml(-1) levels. Protein contents in human serums were analyzed by using the present procedure, and reasonable agreements were obtained with those obtained by a documented spectrophotometric (Biuret) method.

Encapsulation of silica nano-spheres with polymerized ionic liquid for selective isolation of acidic proteins

A nanocomposite is prepared by encapsulating silica nano-spheres with polymerized ionic liquid in aqueous medium without use of any organic solvents. Vinyl groups are covalently introduced on to the surface of silica nano-spheres, which are then encapsulated by copolymerization of 1-vinyl-3-ethylimidazolium bromide (monomer) and 1,4-butanediyl-3,3′-bis-l-vinylimidazolium dibromide (cross-linker) at room temperature. The derived nanocomposite, PIL@SiO2, provides a green adsorbent for protein sorption. PIL@SiO2 is selective toward acidic proteins, and its selectivity can be controlled via varying the amount of monomer used in the copolymerization process. At pHxa06.0, use of 5xa0mg PIL@SiO2 nanocomposite results in a sorption efficiency of up to 95xa0% for 200xa0mgxa0L−1 ovalbumin in 1xa0mL sample solution. Electrostatic and hydrophobic interactions between PIL@SiO2 and protein species dominate the adsorption process. The ovalbumin adsorption behavior is consistent with the Langmuir model, giving a sorption capacity of 333.3xa0mgxa0g−1. The retained ovalbumin is recovered by elution with 0.2xa0% SDS solution. Circular dichroism spectra reveal virtually no change to the α-helix content of ovalbumin after elimination of SDS by use of dialysis. In summary, high-purity ovalbumin is isolated from chicken egg-white by use of the PIL@SiO2 nanocomposite as adsorbent.

Extraction of Cytochrome C by Ionic Liquid 1-Butyl-3-trimethylsilylimidazolium Hexafluorophosphate

A novel method was developed for the extraction of cytochrome c (Cyt-c) from aqueous solution into ionic liquid 1-butyl-3-trimethylsilylimidazolium hexafluorophosphate (BtmsimPF6). At pH 1, an extraction efficiency of ca. 85% was achieved by extracting 3 ml of Cyt-c aqueous solution (5 ng μl−1) with 400 μl of BtmsimPF6 for 30 min. The extraction mechanisms were investigated by UV-Vis and Circular Dichroism (CD) spectra. It was demonstrated that conformation change of Cyt-c was encountered in an acidic medium (pH < 5.0) accompanied by the unfolding of peptide chain and the exposure of hydrophobic groups of heme, which results in the dissolution of Cyt-c in ionic liquid BtmsimPF6. Meanwhile, a cleavage of the sixth coordination bond of the iron atom occurred at pH 1.0 by releasing the Met-80 ligand and giving rise to a vacant coordination position. Thus, the covalent coordination between the cationic Btmsim+ of ionic liquid and the iron atom of heme group facilitates the transfer of Cyt-c into ionic liquid BtmsimPF6.

A spectrophotometric procedure for DNA assay with a micro-sequential injection lab-on-valve meso-fluidic system.

A micro-sequential injection spectrophotometric procedure for DNA assay was developed based on the employment of a lab-on-valve (LOV) meso-fluidic analytical system. A small amount of crystal violet solution (10mul) was de-colored inside the flow cell of the LOV at the presence of 5mul lambda-DNA/HindIII within a certain pH range, and the absorbance decrease of crystal violet solution at 591nm was measured via optical fibers and was employed as the basis of quantification. A uni-variant approach was adopted for the optimization of experimental parameters, including buffer pH, concentration and volume of crystal violet solution, reaction time and sample/reagent loading flow rates. A linear calibration graph was obtained within 0.2-6.0mugml(-1), along with a detection limit of 0.07mugml(-1). The procedure was applied for the determination of lambda-DNA/HindIII in synthetic samples in comparison with a documented procedure.

A novel flow-through fluorescence optosensor for the sensitive determination of tetracycline

A flow-through fluorescence optosensor with Sephadex G-50 microbeads as solid support is developed for the sensitive determination of tetracycline (TC). The fluorescent TC derivative encapsulated in CTAB micelle structures is retained onto the surface of the microbeads packed into a fluorescent flow cell in a flow system, followed by measurement of the native fluorescence of the TC derivative on the bead surface. The retained TC derivative is easily stripped off with DI water from the bead surface by breaking-down the micelle structure. This offers a convenient and effective way for the regeneration of the used solid support with DI water as a carrier. Under the optimal conditions, a linear calibration graph is obtained within a range of 3-500 μg L(-1), along with a detection limit of 1.0 μg L(-1). The present solid surface fluorescence optosensor provides a 22-fold improvement on the detection sensitivity for TC in comparison with that derived by fluorescence detection in aqueous medium. The feasibility of this flow-through fluorescence optosensor is evaluated by analyzing TC in a commercial drug tablet and surface water samples.

Selective isolation of hemoglobin by use of imidazolium-modified polystyrene as extractant

AbstractIonic liquids have attracted much attention in the analysis of a variety of species. The functional groups in ionic liquids can result in highly efficient separation and enrichment and, because of their typical lack of volatility, they are environmentally benign. We grafted imidazole cations onto the surface of chloromethyl polystyrene, denoted PS-CH2-[MIM]+Cl−, and this modified polymer was used to selectively extract the protein hemoglobin (Hb). The prepared extractant PS-CH2-[MIM]+Cl−, containing 2xa0mmol immobilized imidazole groups per gram polymer, was characterized by FT-IR, surface charge analysis, and elemental analysis. The adsorption efficiency was 91xa0%. The adsorption capacity of the PS-CH2-[MIM]+Cl− for Hb was 23.6xa0μgxa0mg−1, and 80xa0% of the retained Hb could be readily recovered by use of 0.5xa0% (m/v) aqueous sodium dodecyl sulfate (SDS) solution as eluate. The activity of the eluted Hb was approximately 90xa0%. The prepared imidazole-containing solid phase polymer was used for direct adsorption of Hb without use of any other solid matrix as support of the ionic liquid. The material was used in practice to isolate Hb from human whole blood.n FigureCoordination interaction between heme of hemoglobin and imidazolium-modified chloromethyl polystyrene.

Determination of Proteins in a Mesofluidic Lab-on-Valve System

Abstract The application of a mesofluidic lab-on-valve system to the spectrophotometric determination of protein was investigated. Protein species in the sample solution reacts rapidly with Congo red at pH 4.1, which forms a complex with a maximum absorption at 496 nm. A univariant approach was used for the optimization of the experimental parameters. A sample volume of 20 μl was used along with 4.0 μl of Congo red solution of 0.9 g l−1, and a flow rate for the detection process of 20 μl s−1 was used. Under optimal conditions, a linear calibration curve was obtained in the range of 12.5–200 μg ml−1 of bovine serum album, along with a detection limit (3σ) of 5.6 μg ml−1 and a sampling frequency of 60 per hour. Protein concentrations in human serums, urine, milk, and yoghourt were determined using this procedure, and satisfactory agreements were obtained with that achieved using the Coomassie brilliant blue method.