Surface Of Silica Beads Research Articles

Temperature-modulated antibody drug separation using thermoresponsive mixed polymer brush-modified stationary phase

Recently, antibody drugs have been investigated for medical treatments of intractable diseases. An effective antibody drug separation method is in demand for production of such drugs. We developed an antibody drug separation material using a mixed polymer brush composed of poly(N-isopropylacrylamide) (PNIPAAm), which is a thermoresponsive polymer, and poly(4-vinylpyridine) (P4VP), which is an antibody-affinity polymer, for temperature modulation and adsorption of the antibody drug rituximab that results in its separation. A mixed polymer brush was prepared on silica bead surfaces by radical polymerization to modify P4VP, followed by atom transfer radical polymerization to modify PNIPAAm. The prepared mixed polymer brush was characterized by CHN elemental analysis, chloride analysis, nitrogen adsorption, thermogravimetric analysis, differential thermal analysis, differential scanning calorimetry, Fourier-transform infrared spectroscopy, X-ray photoelectron spectroscopy, scanning electron microscopy, transmission electron microscopy, and zeta potential measurement. Temperature-modulated adsorption of rituximab on the mixed polymer brush was achieved by concealing and exposing P4VP through the extension and shrinkage of PNIPAAm in the brush, and was investigated using mixed polymer brush-modified silica beads as chromatographic packing materials. By utilizing the specific properties of mixed polymer brushes, rituximab and bovine serum albumin could be separated by simply changing the column temperature. Thus, the mixed polymer brush-modified beads prepared in this study may be useful for purifying rituximab by changing the temperature.

U(VI) and Th(IV) recovery using silica beads functionalized with urea- or thiourea-based polymers – Application to ore leachate

Urea and thiourea have been successfully deposited at the surface of silica beads (through one-pot reaction with formaldehyde) for designing new sorbents for U(VI) and Th(IV) recovery (UR/SiO2 and TUR/SiO2 composites, respectively). These materials have been characterized by FTIR, titration, elemental analysis, BET, TGA, SEM-EDX for identification of structural and chemical properties, and interpretation of binding mechanisms. Based on deprotonation of reactive groups (amine, carbonyl, or thiocarbonyl) and metal speciation, the optimum pH was ~4. Uptake kinetics was fast (equilibrium within 60–90 min). Although the kinetic profiles are fitted by the pseudo-first order rate equation, the resistance to intraparticle diffusion cannot be neglected. Sorption isotherms were fitted by Langmuir equation (maximum sorption capacities: 1–1.2 mmol g−1). Thermodynamics are also investigated showing differences between the two types of functionalized groups: exothermic for TUR/SiO2 and endothermic for UR/SiO2. Metal desorption is highly effective using 0.3–0.5 M HCl solutions: total desorption occurs within 30–60 min; sorption/desorption properties are remarkably stable for at least 5 cycles. The sorbents have marked preference for U(VI) and Th(IV) over alkali-earth and base metals at pHeq ~4.8. By preliminary precipitation steps, it is possible “cleaning” ore leachates of pegmatite ore, and recovering U(VI) and Th(IV) using functionalized silica beads. After elution and selective recovery by precipitation with oxalate (Th-cake) and alkaline (U-cake), the metals can be valorized.

Anion species-triggered antibody separation system utilizing a thermo-responsive polymer column under optimized constant temperature

Although the field of antibody drugs has grown larger, the antibody production still faces several challenges. Effective antibody purification is required, but the conventional purification method for antibodies is cost intensive and often causes aggregation problems, indicating the need for new alternative antibody purification methods. In the present study, a constant temperature antibody purification system for use with a thermo-responsive polymer column was developed based on switching of anion species in eluents. By adjusting the temperature for each antibody, the developed column enabled separation of the therapeutic monoclonal antibodies, rituximab and trastuzumab, from contaminants without changing salt concentration or pH of the eluents. The thermo-responsive hydrogel-modified column packing material was synthesized by introducing n-butyl methacrylate, acrylic acid, N,N′-methylenebisacrylamide and N-isopropylacrylamide to the surface of silica beads with an initiator by a graft-from approach. Elution behavior of antibodies with three types of anions, such as citrate, phosphate, and chloride were tested under three different temperature conditions. It was demonstrated that the thermo-responsive hydrogel grafted column showed a switchable antibody retention behavior at constant temperature and salt concentration, with antibody adsorption by NaCl eluent and desorption by citric acid buffer eluent.

Hierarchically Imprinted Polymer for Peptide Tag Recognition Based on an Oriented Surface Epitope Approach.

FLAG tag (DYKDDDDK) is a short peptide commonly used for the purification of recombinant proteins. The high price of the affinity columns and their limited reusability are a shortcoming for their widespread use in biotechnology applications. Molecularly imprinted polymers (MIPs) can circumvent some of the limitations of bioaffinity columns for such applications, including long-term stability, reusability, and cost. We report herein the synthesis of MIPs selective to the FLAG tag by hierarchical imprinting. Using the epitope imprinting approach, a 5-amino acid peptide DYKDC was selected as a template and was covalently immobilized on the surface of microporous silica beads, previously functionalized with different aminosilanes, namely, 3-(2-aminoethylamino)propyldimethoxymethylsilane, AEAPMS, and N-(2-aminoethyl)-2,2,4-trimethyl-1-aza-2-silacyclopentane, AETAZS. We investigated the effect of the type of silane on the production of homogeneous silane-grafted layers with the highest extent of silanol condensation as possible using 29Si CP/MAS NMR. We observed that the right orientation of the imprinted cavities can substantially improve analyte recoveries from the MIP. After template and silica removal, the DYKDC-MIPs were used as sorbents for solid-phase extraction (molecularly imprinted solid-phase extraction) of the FLAG peptide, showing that the polymer prepared with AETAZS-bound silica beads contained binding sites more selective to the tag (RMIP-AZA = 87.4% vs RNIP-AZA = 4.1%, n = 3, RSD ≤ 4.2%) than those prepared using AEAPMS (RMIP-DM = 73.4% vs RNIP-DM = 23.2%, n = 3, RSD ≤ 4.0%) as a functionalization agent. An extensive computational molecular modeling study was also conducted, shedding some light on the interaction mechanism between the FLAG peptide and the imprinted template in the binding cavities.

Purification and Functional Characterization of CD34-Expressing Cell Subsets Following Ex Vivo Expansion of Umbilical Cord Blood-Derived Endothelial Colony-Forming Cells.

Fluorescent-activated cell sorting (FACS) remains a powerful tool to enrich blood-derived progenitor cells for the establishment of highly proliferative endothelial colony-forming cells (ECFC). Further investigation remains necessary to determine whether the retention of progenitor cell phenotypes after expansion can identify ECFC with enhanced proangiogenic and regenerative functions. This study employed FACS purification to segregate umbilical cord blood-derived ECFC using conserved provascular progenitor cell markers CD34 or aldehyde dehydrogenase (ALDH) activity. ECFC FACS purified for high versus low ALDH activity formed single cell-derived colonies and demonstrated tubule formation in Matrigel at comparable rates. Surprisingly, FACS purification of ECFC for CD34 enriched cells with enhanced colony-forming capabilities and tubule formation within the CD34- population. CD34 expression was enriched on early ECFC populations; however, steady-state expression of CD34 rapidly declined and stabilized on expanded ECFC after serial passage. CD34 expression on ECFC was shown to be cell density dependent and coincided with a loss of progenitor cell characteristics in vitro. Silica-bead surface membrane capture followed by proteomic analysis by label-free liquid chromatography tandem mass spectrometry (LC-MS/MS) identified >100 distinctions (P < 0.05) associated with the plasma membrane of CD34- versus CD34+ ECFC, including a significant enrichment of CD143 (angiotensinogen converting enzyme) on CD34+ cells. Despite an enrichment for traditional endothelial cell markers on the CD34+ ECFC in vitro, implantation of both CD34+ and CD34- ECFC within Matrigel plugs in immunodeficient NOD.SCID mice promoted the formation of vessel-like structures with equivalent integration of human cells at 7 days post-transplantation. Although positive selection of CD34 enriched ECFC establishment before culture, FACS-purified CD34+ ECFC demonstrated reduced colony and tubule formation in vitro, yet demonstrated equivalent vessel formative function in vivo compared to CD34- counterparts. The knowledge will support future studies aiming to identify ECFC subsets with enhanced vessel forming functions for applications of regenerative medicine.

Temperature-modulated cell-separation column using temperature-responsive cationic copolymer hydrogel-modified silica beads.

There is strong demand for cell separation methods that do not decrease cell activity or modify cell surfaces. Here, new temperature-modulated cell-separation columns not requiring cell-surface premodification are described. The columns were packed with temperature-responsive cationic polymer hydrogel-modified silica beads. Poly(N-isopropylacrylamide-co-n-butyl methacrylate-co-N,N-dimethylaminopropyl acrylamide) hydrogels with various cationic moieties were attached to silica-bead surfaces by radical polymerization using N,N'-methylenebisacrylamide as a crosslinking agent. The beads were packed into solid-phase extraction columns, and temperature-dependent cell elution from the columns was found using HL-60 and Jurkat cells. The retention HL-60 and Jurkat cells in columns containing cationic beads at 37 °C was 95.3% to 99.6% and 95.0% to 98.8%, respectively. By contrast, beads without cationic properties exhibited low cell retention (20.6% for HL-60 and 32.5% for Jurkat cells). The cells were mainly retained through both electrostatic and hydrophobic interactions. The retained HL-60 (4.9%) and Jurkat cells (40%) were eluted at 4 °C from the column with a low composition of cationic monomer (DMAPAAm, 1 mol% in copolymer), because the temperature-responsive hydrogels on the beads became hydrophilic, decreasing the hydrophobic interactions between the cells and the beads. A higher number of Jurkat cells than HL-60 cells were eluted because of differences in their electrostatic properties (Jurkat cells: -2.53 mV; HL-60 cells: -20.7 mV). The results indicated that cell retention by the hydrogel-coated beads packed in a solid phase extraction column could be modulated simply by changing the temperature.

Preparation and evaluation of maltose modified polymer-silica composite based on cross-linked poly glycidyl methacrylate as high performance liquid chromatography stationary phase

A new maltose modified polymer-silica composite was fabricated and applied as high performance liquid chromatography (HPLC) stationary phase. The cross-linked poly glycidyl methacrylate (pGMA) layer was chemically bonded to the outer surface as well as pore inner surface of silica beads via in-situ polymerization, and then maltose was modified onto the polymer layer via a [3 + 2] “click” reaction. The porous spherical silica (4 μm diameter) with 300 Å pore size was selected as the matrix so that the 3.25 nm-thick polymer layer fabricated on the pore inner surface would not affect its permeability. The typical ‘U-shape’ retention curves indicated a mixed-mode retention mechanism of the as-synthesized stationary phase. Both polar and non-polar analytes could be well separated on the stationary phase with column efficiency reaching 123809 plates/m for guanosine in hydrophilic interaction liquid chromatography (HILIC) mode and 46808 plates/m for fluorene in reversed-phase liquid chromatography (RPLC) mode, respectively. Nucleotides and their bases were baseline separated with good peak shape without any buffer salt in mobile phase, suggesting the effective shielding of the silanol groups. The packing material also showed excellent chromatographic repeatability with intraday RSDs of the retention time of five nucleosides less than 0.048% (n = 3) and interday RSDs less than 0.33% (n = 7) and great pH stability (from 1.5 to 10.2). Finally, the stationary phase was applied to the separation of ginseng extract.

Creating Sub-50 Nm Nanofluidic Junctions in PDMS Microfluidic Chip via Self-Assembly Process of Colloidal Particles.

Polydimethylsiloxane (PDMS) is the prevailing building material to make microfluidic devices due to its ease of molding and bonding as well as its transparency. Due to the softness of the PDMS material, however, it is challenging to use PDMS for building nanochannels. The channels tend to collapse easily during plasma bonding. In this paper, we present an evaporation-driven self-assembly method of silica colloidal nanoparticles to create nanofluidic junctions with sub-50 nm pores between two microchannels. The pore size as well as the surface charge of the nanofluidic junction is tunable simply by changing the colloidal silica bead size and surface functionalization outside of the assembled microfluidic device in a vial before the self-assembly process. Using the self-assembly of nanoparticles with a bead size of 300 nm, 500 nm, and 900 nm, it was possible to fabricate a porous membrane with a pore size of ~45 nm, ~75 nm and ~135 nm, respectively. Under electrical potential, this nanoporous membrane initiated ion concentration polarization (ICP) acting as a cation-selective membrane to concentrate DNA by ~1,700 times within 15 min. This non-lithographic nanofabrication process opens up a new opportunity to build a tunable nanofluidic junction for the study of nanoscale transport processes of ions and molecules inside a PDMS microfluidic chip.

Thermoresponsive anionic block copolymer brushes with a strongly anionic bottom segment for effective interactions with biomolecules

Thermoresponsive anionic block copolymer brushes were prepared on silica bead surfaces by multistep surface-initiated atom-transfer radical polymerization. The anionic properties of the prepared brushes changed with temperature changes.

A novel automated device for rapid nucleic acid extraction utilizing a zigzag motion of magnetic silica beads

We report a novel automated device for nucleic acid extraction, which consists of a mechanical control system and a disposable cassette. The cassette is composed of a bottle, a capillary tube, and a chamber. After sample injection in the bottle, the sample is lysed, and nucleic acids are adsorbed on the surface of magnetic silica beads. These magnetic beads are transported and are vibrated through the washing reagents in the capillary tube under the control of the mechanical control system, and thus, the nucleic acid is purified without centrifugation. The purified nucleic acid is automatically extracted in 3 min for the polymerase chain reaction (PCR). The nucleic acid extraction is dependent on the transport speed and the vibration frequency of the magnetic beads, and optimizing these two parameters provided better PCR efficiency than the conventional manual procedure. There was no difference between the detection limits of our novel device and that of the conventional manual procedure.We have already developed the droplet-PCR machine, which can amplify and detect specific nucleic acids rapidly and automatically. Connecting the droplet-PCR machine to our novel automated extraction device enables PCR analysis within 15 min, and this system can be made available as a point-of-care testing in clinics as well as general hospitals.

Thermoresponsive anionic copolymer brushes containing strong acid moieties for effective separation of basic biomolecules and proteins.

A thermoresponsive copolymer brush possessing the sulfonic acid group, poly(N-isopropylacrylamide (IPAAm)-co-2-acrylamido-2-methylpropanesulfonic acid (AMPS)-co-tert-butylacrylamide (tBAAm)), was grafted onto the surface of silica beads through surface-initiated atom transfer radical polymerization. Prepared copolymer and copolymer brushes on silica beads were characterized by observing the phase transition profile, CHNS elemental analysis, X-ray photoelectron spectroscopy, and gel permeation chromatography. The phase transition profile indicated that an appropriate AMPS composition for enabling thermally modulated property changes is 5 mol %, while excessive amounts of sulfonic acid groups prevented copolymer phase transition. Chromatographic elutions of catecholamine derivatives and basic proteins were observed, using the prepared copolymer brush-modified beads as chromatographic matrices, and the results suggest that the beads interact with these analytes through relatively strong electrostatic interactions. Thus, poly(IPAAm-co-AMPS-co-tBAAm) brush-modified beads will be useful for effective thermoresponsive chromatography matrices that separate basic biomolecules through strong electrostatic interactions.

Preparation of low-density 90Y microspheres consisting of mesoporous silica core/yttria shell: a potential therapeutic agent for hepatic tumors

A new low-density silica core/yttria shell microspheres was developed for medical purposes. For preparing the core of this material, sol-emulsion-gel method was used. It was then calcined at 1,000 °C to increase the strength of the mesoporous beads. After cooling, it was added into a solution containing the mixture of yttrium chloride and urea. Under these circumstances, yttrium chloride can decompose to yttrium basic carbonate particles by the gradual hydrolysis of urea. These particles are able to deposit on the surface of mesoporous silica beads. Different coated samples were prepared under various pH, temperature and stirring conditions. Samples were heated to convert yttrium basic carbonate to yttria. The resulting silica core/yttria shell microspheres was characterized by laser diffractometer, scanning electron and optical microscopy techniques. It was found the pH of silica sols, aging time and Span-80 play important role in the stage of mesoporous bead preparation. Furthermore, the findings showed the heating of core/shell microspheres was essential for the ultimate strength of the shell. The in vitro chemical durability tests showed the release of yttrium ion in two simulated body fluids at pH 6 and 7 was negligible after 4 weeks.

Thermoresponsive Copolymer Brushes Possessing Quaternary Amine Groups for Strong Anion-Exchange Chromatographic Matrices

A thermoresponsive copolymer incorporating a quaternary amine group, poly(N-isopropylacrylamide-co-3-acrylamidopropyl trimethylammonium chloride (APTAC)-co-tert-butylacrylamide), was conjugated to the surface of silica beads through surface-initiated atom transfer radical polymerization. Prepared copolymer- and copolymer brush-modified beads were characterized by CHN elemental analysis, X-ray photoelectron spectroscopy, gel permeation chromatography, and observation of phase transition profiles. Phase transition profiles of the prepared copolymer indicated that 5 mol % APTAC is suitable for enabling thermally modulated property changes in the copolymer. Chromatographic elution behaviors of adenosine nucleotides and proteins were observed using prepared beads as chromatography matrices. Higher retention time of adenosine nucleotides and strong protein adsorption behavior were observed compared with those on beads with tertiary amine groups, because of the strong basic properties. Therefore, copolymer brush modified beads will be useful as thermoresponsive ion-exchange chromatographic matrices.

Single white light emitting hybrid nanoarchitectures based on functionalized quantum dots

Colloidal white emitting nanostructures were successfully fabricated by covalently binding a blue emitting oligofluorene at the surface of silica beads, that incorporate orange luminescent colloidal CdSe@ZnS quantum dots (QDs). White light was achieved by carefully tuning the size of the QDs to complementarily match the emission color of the blue fluorophore and taking into account the delicate balance between the emission of the QDs in the core of the silica beads and the amount of the organic dye bound to the silica surface. The proposed approach is highly versatile as it can be extended to the fabrication of a variety of luminescent hybrid nano-objects, playing with the complementarity of the emission color of the inorganic and organic fluorophores at the nanoscale.

Removal of Endotoxin from Collagen Solution by Lectin-grafted Silica Beads

In the present study,we propose to study endotoxin absorption/removal processes in collagen solutions by taking advantage of the unique adsorption characteristics of lectin molecules specifically onto the poly-saccharide components in endotoxin.The surface of silica beads was pretreated with acid-etching process leaving reactive OH groups which facilitated the adoption of amino-siloxane group for grafting of lectin molecules.The preliminary results demonstrated that high levels of adsorption of endotoxin were all readily achievable(99%) under grafting formulation with various concentrations of lectin,such as 1,2 and 5 g/L.The results also indicated that 30 pieces of lectin-grafted silica beads would efficiently adsorb endotoxin from a volume of 9.6 mL collagen solution.For future work,we plan to evaluate designs of column adsorption method and system for the removal of endotoxin in collagen material.

Protein–polyphenol interaction on silica beads for astringency tests based on eye, photography or reflectance detection modes

Astringency is an organoleptic property of beverages and food products resulting mainly from the interaction of salivary proteins with dietary polyphenols. It is of great importance to consumers, but the only effective way of measuring it involves trained sensorial panellists, providing subjective and expensive responses. Concurrent chemical evaluations try to screen food astringency, by means of polyphenol and protein precipitation procedures, but these are far from the real human astringency sensation where not all polyphenol–protein interactions lead to the occurrence of precipitate. Here, a novel chemical approach that tries to mimic protein–polyphenol interactions in the mouth is presented to evaluate astringency. A protein, acting as a salivary protein, is attached to a solid support to which the polyphenol binds (just as happens when drinking wine), with subsequent colour alteration that is fully independent from the occurrence of precipitate. Employing this simple concept, Bovine Serum Albumin (BSA) was selected as the model salivary protein and used to cover the surface of silica beads. Tannic Acid (TA), employed as the model polyphenol, was allowed to interact with the BSA on the silica support and its adsorption to the protein was detected by reaction with Fe(III) and subsequent colour development. Quantitative data of TA in the samples were extracted by colorimetric or reflectance studies over the solid materials. The analysis was done by taking a regular picture with a digital camera, opening the image file in common software and extracting the colour coordinates from HSL (Hue, Saturation, Lightness) and RGB (Red, Green, Blue) colour model systems; linear ranges were observed from 10.6 to 106.0 μmol L−1. The latter was based on the Kubelka–Munk response, showing a linear gain with concentrations from 0.3 to 10.5 μmol L−1. In either of these two approaches, semi-quantitative estimation of TA was enabled by direct eye comparison. The correlation between the levels of adsorbed TA and the astringency of beverages was tested by using the assay to check the astringency of wines and comparing these to the response of sensorial panellists. Results of the two methods correlated well. The proposed sensor has significant potential as a robust tool for the quantitative/semi-quantitative evaluation of astringency in wine.

Tether-supported biomembranes with α-helical peptide-based anchoring constructs.

The strict requirement of constructing a native lipid environment to preserve the structure and functionality of membrane proteins is the starting constraint when building biomaterials and sensor systems from these biomolecules. To enhance the viability of supported biomembranes systems and build new ligand display interfaces, we apply rationally designed peptides partitioned into the lipid bilayer interface. Peptides designed to form membrane-spanning α-helical anchoring domains are synthesized using solid-phase peptide synthesis. K(3)A(4)L(2)A(7)L(2)A(3)K(2)-FITC is synthesized on the 100 mg scale for use as a biomembrane anchoring molecule, where orthogonal side-chain modifications allow us to introduce probes enabling peptide localization within supported bilayers. The peptides are found to form α-helical domains within liposomes as assessed with circular dichroism spectroscopy. These peptides are designed to be incorporated into lipid bilayers supported by microspheres and serve as biomembrane anchoring moieties to amino-terminated surfaces. Here, the silica bead surface (4.7 μm diameter) is activated with homobifunctional NHS-PEG(3000)-NHS as "polymer cushion" spacers. This tethering to a subset of the K(3)A(4)L(2)A(7)L(2)A(3)K(2)-FITC molecules present in the bilayer is achieved by the fusion of liposomes followed by coupling of the peptide amino groups to the NHS presented from the silica microsphere surfaces. The biomembrane distributions of tethered and untethered K(3)A(4)L(2)A(7)L(2)A(3)K(2)-FITC are probed with confocal microscopy and are found to give 3D reconstructions consistent with largely homogeneous supported biomembranes. The fluidity of the untethered fraction of peptides within supported membranes is quantified using the fluorescence recovery after photobleaching (FRAP) technique. The presence of the PEG(3000) polymer cushion facilitated a 28.9% increase in peptide diffusivity over untethered bilayers at the lowest peptide to lipid ratio we examined. We show that rationally designed peptide-based anchors can be used to tether lipid bilayers, creating a polymer-cushioned lipid microenvironment on surfaces with high lateral mobility and facilitating the development of a new platform for ligand displays.

Poly(N-isopropylacrylamide)-modified silica beads with hyperbranched polysiloxysilane for three-dimensional cell cultivation

The immobilization of poly(N-isopropylacrylamide) (PNIPAM) onto the surface of silica beads via hyperbranched polysiloxysilane (HBPS) was studied to develop a new material for cell cultivation. HBPS terminated with a vinyl functional group was synthesized by self-polymerization of an AB2 monomer. The terminal vinyl group was converted into a chain-transfer agent for reversible addition fragmentation chain transfer (RAFT) polymerization. The obtained HBPS was immobilized on the silica surface by mixing silica beads and HBPS in hexane. The graft polymerization of PNIPAM onto the silica surface was achieved using RAFT polymerization. The molecular weights of the grafted polymers obtained by cleavage from the surface of silica beads using NaOH aqueous solution were in the range of 6000–23 000 and the polydispersity index values were 1.4–1.6. The grafted silica beads have been tested for cell cultivation. Silica-HBPS-g-PNIPAM with a higher molecular weight (Mn) of PNIPAM (cleaved PNIPAM Mn=14 800 and 23 700) showed a thermo-reversible phase transition at approximately 34 °C, and controlled cell adhesion and detachment was demonstrated with these beads.

High Stability of Thermoresponsive Polymer-Brush-Grafted Silica Beads as Chromatography Matrices

Thermo-responsive chromatography matrices with three types of graft architecture were prepared, and their separation performance and stability for continuous use were investigated. Poly(N-isopropylacrylamide)(PIPAAm) hydrogel-modified silica beads were prepared by a radical polymerization through modified 4,4'-azobis(4-cyanovaleric acid) and N,N'-methylenebisacrylamide. Dense PIPAAm brush-grafted silica beads and dense poly(N-tert-Butylacrylamide (tBAAm)-b-IPAAm) brush-grafted silica beads were prepared through a surface-initiated atom transfer radical polymerization (ATRP) using CuCl/CuCl(2)/ Tris(2-(N,N-dimethylamino)ethyl)amine (Me(6)TREN) as an ATRP catalytic system and 2-propanol as a reaction solvent. Dense PIPAAm brush-grafted silica beads exhibited the highest separation performance because of their strong hydrophobic interaction between the densely grafted well-defined PIPAAm brush on silica-bead surfaces and analytes. Using an alkaline mobile phase, dense themoresponsive polymer brushes, especially having a hydrophobic basal layer, exhibited a high stability for continuous use, because polymer brush on the silica bead surfaces prevented the access of water to silica surface, leading to the hydrolysis of silica and cleavage of grafted polymers. Thus, the precisely modulating graft configuration of thermoresponsive polymers provided chromatography matrices with a high separation efficiency and stability for continuous use, resulting in elongating the longevity of chromatographic column.

Preparation of Composites of Silica/PNIPAm by Coupling Reaction and Their Application in HPLC

Poly(N-isopropylacrylamide) was grafted on the surface of ultrafine silica beads through coupling reaction of semi-telechelic poly(N-isopropylacrylamide) (PNIPAm) and the silica beads. The synthetic route included the preparation of semi-telechelic PNIPAm with triethoxysilan group at the polymer chain end and the coupling reaction of the semi-telechelic PNIPAm with silica particles. The characterizations by infrared spectrometry, differential scanning calorimetry, scanning electron microscopy, and thermogravimetry analysis confirmed the formation of core-shell composites of silica-PNIPAm. The composites were thermo-sensitive and were used as a packing material in high-performance liquid chromatography (HPLC) for separating organic compound mixtures. The result indicated that satisfactory separations could be achieved by controlling the column temperature.