Ind. Eng. Chem. Res. 2007, 46, 2117-2124
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Surface-Grafted, Molecularly Imprinted Polymers Grown from Silica Gel for Chromatographic Separations Xiaolin Wei and Scott M. Husson* Department of Chemical and Biomolecular Engineering, Clemson UniVersity, Clemson, South Carolina 29634-0909
Imprinted polymers were grafted successfully from silica gel via atom-transfer radical polymerization. Measured binding kinetics for protected amino acid templates onto the molecularly imprinted polymers on silica gel (MIP-SG) and MIPs prepared by conventional solution-phase synthesis demonstrated that the MIP-SG had improved mass-transfer properties and also had higher binding capacity per unit mass of polymer. In addition, a high-performance liquid chromatography column packed with MIP-SG showed higher column efficiency and better resolution for the enantiomers, Boc-L-tryptophan and Boc-D-tryptophan, relative to a column packed with the conventionally prepared MIP. When Boc-L-Trp was used as the template molecule during MIP-SG synthesis, the column packed with the resulting polymer retained the Boc-L-Trp to longer elution time than the Boc-D-Trp. This order of elution was switched when Boc-D-Trp was used as template. Introduction Molecularly imprinted polymers (MIPs) have gained increasing attention over the past 3 decades for a diverse range of applications such as optical resolution of chiral materials;1 separation or enrichment of drugs,2 pesticides and herbicides;3 mimicking antibody and receptor binding sites in recognition and assay systems,4 and catalytic enzyme sites;5 and chemical and biochemical sensing.6 For these and other applications, an MIP material would perform ideally if it was limited only by the intrinsic kinetics of association of the template molecule with imprint sites and not by diffusional mass transfer within the MIP phase. Generally speaking, however, the conventional methods that are used lead to MIP materials whose performances are limited by slow intraparticle mass transfer and nonquantitative template recovery. In recent years, to overcome these drawbacks, a new molecular imprinting genre called “surface molecular imprinting” or 2-D imprinting has emerged. Surface imprinting is in its early stages, and several methods have been described by our group and others. Perez et al.7 have developed a surface imprinting technique based on emulsion polymerization in which small beads are generated in an oil-in-water system in the presence of surfactant, and the template molecule is part of the surfactant that resides at the two-phase interface. Mosbach and colleagues8 introduced a protocol to create surface binding sites by immobilizing the template molecule on porous silica beads prior to polymerization, followed by dissolution of the silica after polymerization. MIPs also have been synthesized on the surface of silica gel9 and capillary columns10,11 by using chemically bound initiators, on the surface of polymeric membranes via physically absorbed initiators,12 from the surface of gold electrodes by electropolymerization13,14 and surfaceconfined atom-transfer radical polymerization,15,16 from the surface of polyacrylonitrile membranes by photopolymerization,17,18 and on the surface of ultrathin titania gel films by a surface sol-gel process.19 The approaches9,20,21 to graft MIPs from silica or resin surfaces using photopolymerization with a surface-bound photoiniferter or reversible addition-fragmenta* To whom correspondence should be addressed. E-mail: shusson@ clemson.edu. Tel.: +1 (864) 656-4502. Fax: +1 (864) 656-0784.
tion chain-transfer (RAFT) polymerization with a surface-bound RAFT initiator provide chromatographic materials with improved kinetic properties when compared to the MIPs prepared by the conventional bulk or solution polymerization. Of special interest for chromatographic separations, the modification of silica by polymer coatings or other functional groups has been developed and applied in the chemical process industry,22 environmental science,23 and pharmaceutical24 and agricultural industries,25 as well as in general analytical methods.26 Grafting reactions have been studied extensively in order to tailor the surface properties for desired applications.27,28 Silica can be prepared in various forms with different particle diameters, pore sizes, and pore volumes.29-31 The ease of derivatization of silanol groups via reaction with alkoxysilanes or chlorosilanes makes silica a widely accepted substrate in separation science, especially in chromatography where a large number of “bonded-phase” silicas have been synthesized and commercialized.32,33 Polymer-modified silicas have been used for chiral separation,34 ion separation,35 and protein adsorption36 to name a few applications. This paper describes the use of atom-transfer radical polymerization (ATRP), a catalyst-activated, controlled polymerization technique, to functionalize silica gel with molecularly imprinted polymer films. It also describes the characterization of the physical, chemical, and performance properties of these materials for chromatographic separations. The methodology was extended from our previous work on graft polymerization of imprinted films from flat, gold surfaces.15 The primary goals of this contribution were to transfer the surface imprinting protocol that we developed for gold surfaces to silica gel and to compare the performance of molecularly imprinted polymers on silica gel (MIP-SG) to MIPs prepared by a conventional solution-phase synthesis protocol. Materials and Methods Materials. All reported purities are given in weight percent. 2-Vinylpyridine (2-Vpy, 97%) and ethylene glycol dimethacrylate (EGDMA, 98%) were obtained from Aldrich and purified by passing through a column of activated aluminum oxide (basic, Aldrich), followed by distillation under vacuum at 330 Pa prior to use. The following were used as received: 3-Ami-
10.1021/ie0606284 CCC: $37.00 © 2007 American Chemical Society Published on Web 02/20/2007
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Ind. Eng. Chem. Res., Vol. 46, No. 7, 2007
nopropyldimethylethoxysilane (3-APDMES, 97%) was purchased from Gelest, Inc. (Morrisville, PA); (4-chloromethyl)benzoyl chloride (97%), copper(I) bromide (99.995+%), copper(II) bromide (99.999%), potassium bromide (FT-IR grade, >99%), toluene (anhydrous, 99.8%), N,N-dimethylformamide (DMF, 99%), acetonitrile (HPLC grade, 99.9%), methanol (HPLC grade, 99.9%), and acetone (HPLC grade, 99.8%) were purchased from Aldrich. Acetic acid (glacial, 99.7%+) and 1,4,8,11-tetramethyl-1,4,8,11-tetraazacyclotetradecane (Me4Cyclam, 97%) were obtained from Alfa Aesar (Ward Hill, MA). Aqueous hydrogen peroxide (35%), hydrochloric acid (37%), Boc-L-Trp (99.8%) and Boc-D-Trp (99.8%) were purchased from Fisher Scientific. Silica gel was kindly provided by Grace GmbH & Co. KG (Worms, Germany); it has irregular shape with average particle size of 20 µm, average pore diameter of 0.1 µm, and surface area and pore volume of 40 m2/g and 1.05 mL/g. Pretreatment and Silanization of Silica Surface. To eliminate any surface contaminants and activate the surface silanol groups for silanization, the silica gel was pretreated. Silica gel (20 g) was suspended in 200 mL of 35% aqueous hydrogen peroxide, and the suspension was stirred thoroughly for 4 h. After activation, the silica was filtered and rinsed with a large volume of deionized water and then dried at 80 °C overnight. The dried silica gel was stored in 200 mL of anhydrous toluene under a nitrogen atmosphere. Silanization is the most common and effective method for modification of silica substrates. 3-APDMES is used widely as a reagent for silanization because its amine group is amenable to further coupling reactions. Di- or trifunctional silane reagents were avoided for this investigation because they can form multiple layers by hydrolysis during silanization;37 this hydrolysis is difficult to control, and, most importantly, thick multilayers may block the inner pores of the silica gel and cause transport problems in the subsequent chromatography application. For silanization, 3-APDMES (3.95 g) was added into the silica gel suspension in 200 mL of toluene, and the suspension was stirred and refluxed at 110 °C for 24 h. The modified silica was filtered, dried at 120 °C for 3 h, washed with 500 mL of methanol, and redried at 50 °C overnight under vacuum at 330 Pa. Anchoring the (4-Chloromethyl)benzoyl Chloride Initiator. Well-dried, amine-functionalized silica (20 g) was contacted with (4-chloromethyl)benzoyl chloride (3.12 g) in 200 mL of DMF for 18 h at room temperature. The reaction of the acid chloride group of (4-chloromethyl)benzoyl chloride with the primary amine group of surface-bound APDMES forms a covalently anchored (4-chloromethyl)benzamide initiator functional group. A constant nitrogen gas purge was used to remove byproduct HCl. This initiator-functionalized silica gel was filtered, washed exhaustively with DMF, and dried overnight at 50 °C under vacuum at 330 Pa. Graft Polymerization of Molecularly Imprinted Polymer from Silica Gel (MIP-SG) via ATRP. The (4-chloromethyl)benzamide initiator groups can be activated by atom-transfer reactions with a suitable catalyst system to form radicals; a typical catalyst comprises copper salts and amine-containing organic ligands. By adjusting the molar ratio of Cu(I) to Cu(II) in the system, the reversible equilibrium between the radical and dormant species can be tuned to give a low instantaneous number (surface concentration) of growing radicals, which, along with fast initiation and negligible irreversible termination and chain-transfer reactions, makes the polymerization proceed in a controllable manner. For this study, CuIBr and CuIIBr2 were
Scheme 1. Chemistries Used To Covalently Bond the Initiator (4-chloromethyl)benzoyl Chloride onto the Silica Gel Surfacea
a The first step (top) uses APDMES to functionalize the surface with reactive amine groups. In a second step (bottom), the acid chloride group of the initiator reacts with the surface amine to tether it to the surface.
used with Me4Cyclam as the ligand to form the organometallic catalyst. 2-Vpy was used as the monomer and EGDMA as the cross-linking agent; acetonitrile was the solvent. A typical reaction procedure follows to prepare the MIPSG: 304 mg (1 mmol) of the template molecule n-Boc-L-Trp was dissolved in a mixture of 1.262 g (12 mmol) of 2-Vpy, 7.929 g (40 mmol) of EGDMA, 2 g of initiator-functionalized silica gel, and enough acetonitrile to make a suspension with a total volume of 20 mL. The suspension was stirred for 30 min and degassed with three freeze-pump-thaw cycles to remove oxygen. Next, the suspension was transferred into a water-free (
