Novel biporous polymeric stationary phase for high-speed protein chromatography

Rigid biporous beads (BiPB) were fabricated by double emulsification. An aqueous suspension of superfine calcium carbonate granules and organic solvent were used as porogenic agents to create superpores and micropores, respectively. The polymerization of monomers, glycidyl methacrylate, and ethylene glycol dimethacrylate was initiated with benzoin ethyl ether by ultraviolet irradiation. Modified with diethylamine (DEA), the BiPB were derivatized into an anion-exchange medium (which is denoted as DEA–BiPB). The DEA–BiPB with an average diameter of 46.3 μm was characterized to possess two types of pores, that is, micropores (20–200 nm) and superpores (500–5300 nm). Flow hydrodynamic experiments showed that the DEA–BiPB column had a smaller backpressure than that of the conventional microporous beads column at a given flow rate. The static adsorption capacity of the DEA–BiPB was close to that of the DEA–MiPB for bovine serum albumin. However, frontal analysis demonstrated that the dynamic binding capacity of the DEA–BiPB column was two times higher than that of the DEA–MiPB at a flow rate of 1800 cm/h. Moreover, the purification of the molecular chaperone GroEL was carried out with the DEA–BiPB column at two flow rates (150 and 1500 cm/h). This showed that the GroEL purification was nearly the same at the two flow rates tested. These results indicate that the DEA–BiPB column is promising for high-speed protein chromatography. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 103: 17–23, 2007

In order to improve Cu(II) adsorption ability of chelating resins based on a macrocyclic ligand 1,4,8,11-tetraazacyclotetradecane-5,7-dione (L) bound to crosslinked copolymers, the resins were prepared by reaction of L with poly(glycidyl methacrylate)s crosslinked with ethylene glycol dimethacrylate (1G), tetraethylene glycol dimethacrylate (4G) or nonaethylene glycol dimethacrylate (9G). By systematically changing the amounts of each crosslinker (degree of crosslinking) and of isobutyl acetate (IBA) as diluent in the suspension polymerization of the monomers, precursory copolymers with various degrees of crosslinking as well as with different specific surface areas (SSAs) were prepared, and then L was introduced into them. Ligand contents, SSAs, and Cu(II) adsorption abilities of the resulting resins were evaluated, and it was clarified that resins derived from precursors (prepared by using 4G or 9G in the presence of 120–160 vol % of IBA) exhibit high capacities for Cu(II), whereas the capacities of resins crosslinked with 1G are not so high. Further detailed study on the uptake of metal ions using the resins RG4D(10)-140 and RG9D(10)-140 (derived from the precursors prepared by using 10 mol % of the respective crosslinkers 4G and 9G in the presence of 140 vol % of IBA) showed that RG9D(10)-140 is much superior to RG4D(10)-140 in the column-mode adsorption of Cu(II). These results indicate that not only the selection of the suitable crosslinker but also the use of a pertinent amount of the diluent are very important in improving the complexing abilities of the polymer-bound L. © 1996 John Wiley & Sons, Inc.

Capillary electrochromatography with macroporous particles

The aim of this work was the preparation of molecularly imprinted membranes by surface modification of microfiltration polyvinylidene fluoride membranes with a thin layer of molecular imprinted polymer by UV irradiation. The flavonoid naringin (4,5,7-trihydroxyflavanone-7-rhamnoglucoside) was used as template molecule. The compound 4-vinylpyridine was used as functional monomer. Benzoin ethyl ether and ethylene glycol dimethacrylate were used as photo-initiator and cross-linker, respectively. Membranes were also modified in the absence of naringin and used as reference (blank). Imprinted membranes prepared in this work could be applied as new separation devices in affinity filtrations aimed to reduce the naringin content in grapefruit and improve its benefit properties. The method used to prepare them offers the advantage of combining the mechanical integrity of the membrane support with the selective recognition properties of the imprinted polymer layer. The recognition properties of all prepared membranes were evaluated by their capability to retain naringin in aqueous solution during filtration tests. Results showed that imprinted membranes exhibited specific binding properties for template molecules. Blank membranes only showed non-specific binding. The selectivity of the naringin-imprinted membranes was tested evaluating their bound property toward the rutin, a structural analogue of naringin.

Crosslinking reaction of polymer by ultraviolet (UV) irradiation has been important in industries. In this work, photocrosslinking of styrene–isoprene–styrene (SIS) triblock copolymer in the presence of benzoin photoinitiator and a dimethacrylate monomer as crosslinking agent was investigated. Curing of samples was initiated under UV irradiation. Benzoin was used as photoinitiator because it contains chromophore group that could absorb UV irradiation. Ethylene glycol dimethacrylate (EGDMA) was used as crosslinking agent, since it has alkene functional groups that could react with the alkene group of SIS. ATR-FTIR spectra of samples show that absorption band of double bond at 1500–1600 cm−1 decreases after UV exposure. Increasing the concentration of benzoin (0.1–1 phr) and EGDMA (1–10 phr) leads to an increase in gel content and hardness, while swelling ratio decreases. After 5 min heating at 150 °C, about 20%wt of the unirradiated compound became insoluble, because heating of compound at 150 °C causes crosslinking reaction without any irradiation.

A polyHIPE is a cross-linked, highly porous polymer from the polymerization of monomers and cross-linking comonomers in the external phase of a high internal phase emulsion (HIPE). Crystallizable polyHIPE were synthesized through the copolymerization of stearyl acrylate (A18) with ethylene glycol dimethacrylate (EGDMA). PolyHIPE synthesized with the more flexible EGDMA were compared to those synthesized with the more rigid divinylbenzene. The polyHIPE densities were around 0.11 g cm-3. EGDMA destabilized the HIPE yielding relatively large voids and a structure whose partial closed-cell nature increased with increasing EGDMA content. The EGDMA cross-linked polyHIPE exhibited higher melting peak temperatures and higher crystallinities than the divinylbenzene cross-linked polyHIPE. Organic phase initiation of the EGDMA cross-linked polyHIPE yielded lower melting temperatures, lower crystallinities, and a highly interconnected porous structure that resulted in a lower modulus.

During reservoir evaluation, the microscopic pore structure of low-rank coal is mainly characterized in order to study the coalbed methane diffuse and migration mechanisms and control. The low-rank coals are very different in pore type and size, so it is necessary to use various techniques to describe their pore structure. For vitrain and durain of the Coal Member of the Yan’an Formation from Huanglong Coalfield, their chemical composition and microscopic pore structure characteristics were studied, and the factors of influencing the pore size distribution (PSD) were explored. Obviously, vitrain and durain are different in chemical composition. Vitrain has higher moisture content, volatile yield, and vitrinite group content than durain. Vitrain and durain mainly contain vitrinite and inertinite, respectively. The pore structure characteristics (e.g., pore types and PSD) of vitrain and durain were systematically by mercury intrusion porosimetry (MIP), low-temperature nitrogen adsorption, and carbon dioxide (CO2) adsorption. The vitrain and durain samples with a micropore size of <2 nm were mainly tested on their specific surface area (SSA) and pore volume (PV). The results show that microporous vitrain has larger SSA and PV than microporous durain, while mesoporous and macroporous vitrain has smaller SSA and PV than mesoporous and macroporous durain. SSA is very positively correlated with PV. The ash content is negatively correlated with SSA and PV. The ash content influences microporous vitrain more greatly than microporous durain, but mesoporous and macroporous durain more greatly than mesoporous and macroporous vitrain. SSA is positively correlated with the vitrinite content of durain and negatively correlated with the inertinite and exinite contents of durain. However, SSA is negatively correlated with the vitrinite and exinite contents of vitrain and positively correlated with the inertinite content of vitrain. Vitrain has higher methane adsorption capacity, desorption rate, and recovery ratio than durain. There are parameters that are obviously affected by the micropore characteristics.

Functional crosslinked polymer particles synthesized by precipitation polymerization for liquid chromatography

Surface modification of macroporous poly(glycidyl methacrylate- co-ethylene glycol dimethacrylate) resins for improved Candida antarctica lipase B immobilization

Membrane-modification effects, induced by ultraviolet (UV) irradiation in diacetylenic liposomes, were analyzed upon contact with cells, biological membranes, and proteins. Liposomes formulated with mixtures of unsaturated 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine and saturated 1,2-dimyristoyl-sn-glycero-3-phosphocholine, in a 1:1 molar ratio, were compared with those that were UV-irradiated and analyzed in several aspects. Membrane polymerization inherence on size stability was studied as well as its impact on mitochondrial and microsomal membrane peroxidation induction, hemolytic activity, and cell viability. Moreover, in order to gain insight about the possible irradiation effect on interfacial membrane properties, interaction with bovine serum albumin (BSA), lysozyme (Lyso), and apolipoprotein (apoA-I) was studied. Improved size stability was found for polymerized liposomes after a period of 30 days at 4°C. In addition, membrane irradiation had no marked effect on cell viability, hemolysis, or induction of microsomal and mitochondrial membrane peroxidation. Interfacial membrane characteristics were found to be altered after polymerization, since a differential protein binding for polymerized or nonpolymerized membranes was observed for BSA and Lyso, but not for apoA-I. The substantial contribution of this work is the finding that even when maintaining the same lipid composition, changes induced by UV irradiation are sufficient to increase size stability and establish differences in protein binding, in particular, reducing the amount of bound Lyso and BSA, without increasing formulation cytotoxicity. This work aimed at showing that the usage of diacetylenic lipids and UV modification of membrane interfacial properties should be strategies to be taken into consideration when designing new delivery systems.

Preparation of hydrogels via ultrasonic polymerization

Controlled protein adsorption on PMOXA/PAA based coatings by thermally induced immobilization

The free radical copolymerization of glycidyl methacrylate (GMA) and octadecyl acrylate (ODA) was carried out at 70 °C in solution using toluene as solvent and 2,2′-azobisisobutyronitrile (AIBN) as initiator. The copolymer compositions were determined using 1H NMR spectroscopy. The monomer reactivity ratios were computed using five linear methods at low conversions, i.e. Mayo–Lewis (ML), Fineman–Ross (FR), inverted Fineman-Ross (IFR), Ezrielev–Brokhina– Roskin (EBR) and Kelen–Tudos (KT). The reactivity ratios resulted from different methods are in agreement with each other and showed higher reactivity for GMA monomer in comparison with ODA monomer (FR: r G = 1.29 and r O = 0.68). Crosslinked GMA–ODA copolymers were also prepared using ethylene glycol dimethacrylate as a crosslinker. Thermal properties and swelling behaviors of crosslinked copolymers in different polar and nonpolar solvents were evaluated. GMA incorporation in the crosslinked copolymers improved T g values, while decreased T m quantities and swelling in nonpolar solvents. The swelling values are in the acceptable range and the prepared functional crosslinked copolymers can be used for nonpolar solvents absorbency applications.

Gradient elution separation and peak capacity of columns packed with porous shell particles

In this work, open-tubular capillary electrochromatography (OT-CEC) method with bare gold nanoparticles (GNPs)-based stationary phase has been developed and applied for separation of tryptic peptide fragments of native and glycated proteins, bovine serum albumin (BSA), and human transferrin (HTF). The GNPs-based stationary phase was prepared by immobilization of bare GNPs, freshly reduced from tetrachloroaurate(III) ions by citrate reduction, on the sol-gel pretreated inner wall of the fused silica capillary. The separation efficiency, peak capacity, and peptide recovery of this open-tubular capillary column were investigated by varying the experimental parameters such as type and concentration of the buffering constituent and pH of the background electrolyte (BGE), temperature, and separation voltage. The best separations of the above tryptic peptides were achieved in the BGE composed of aqueous 100 mmol/L sodium phosphate buffer, pH 2.5, at separation voltage 10 kV per 47-cm long, 50 μm inside diameter capillary thermostated at 25°C. OT-CEC with bare GNPs stationary phase is shown to be a suitable technique for separation of complex peptide mixtures arising from tryptic digestion of native and glycated BSA and HTF, and for investigation of glycation (nonenzymatic glycosylation) of these proteins.