Atom-Transfer Radical Graft Polymerization Initiated Directly from

Initiation of atom-transfer radical polymerization of a number of monomers ..... Journal of Applied Polymer Science 2008 107 (10.1002/app.v107:4), 264...
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Anal. Chem. 2006, 78, 7098-7103

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Atom-Transfer Radical Graft Polymerization Initiated Directly from Silica Applied to Functionalization of Stationary Phases for High-Performance Liquid Chromatography in the Hydrophilic Interaction Chromatography Mode Petrus Hemstro 1 m, Michal Szumski,† and Knut Irgum*

Department of Chemistry, Umeå University, S-90187 Umeå, Sweden

Initiation of atom-transfer radical polymerization of a number of monomers (styrene, methyl acrylate, 3-[N,Ndimethyl-N-(methacryloyloxyethyl)ammonium] propanesulfonate, butyl methacrylate, 2,3-epoxypropyl methacrylate) directly from chlorinated porous silica particles has been performed. The grafting has been confirmed and evaluated by Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. This initiation technique results in a hydrolytically stable initial Si-C bond, tethering the polymer to the silica substrate. The resulting grafted particles have been used as separation materials for both reversed-phase and hydrophilic interaction chromatography. Stationary phases prepared by surface modification of porous silica with organic interactive layers are the foundation of modern liquid chromatography.1 Preparation of bonded stationary phases based on silica has been carried out almost exclusively by the “grafting to” technique, by attachment of alkyl chains (typically octadecyl) to porous silica particles for HPLC applications through silane coupling reactions. However, the attachment of preformed polymers onto surfaces often leads to low grafting density and low film thickness, as the functional moieties must diffuse through the existing polymer film to reach the reactive sites on the surface.2 A more serious weakness with this approach is the thermally and hydrolytically labile Si-O-C bond that links the polymer and the silica support when attachment is established through a silanol condensation reaction. Hydrolysis of this bond leads to ligand stripping under acidic conditions,1 a problem that has been * To whom correspondence should be addressed. Phone: +46-90-7865997. E-mail: [email protected]. † Permanent address: Department of Environmental Chemistry and Ecoanalytics, Faculty of Chemistry, Nicolas Copernicus University, ul. Gagarina 7, 87100 Torun, Poland. (1) Unger, K. K. Porous Silica, Its Properties and Use as Support in Column Liquid Chromatography; Journal of Chromatogr. Library Vol. 16; Elsevier: Amsterdam, 1979. (2) Zajac, R.; Chakrabarti, A. Phys. Rev. E 1995, 52, 6536-6549.

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addressed by using silanes with more than one activated group in combination with a controlled amount of water. This process leads to “polymeric” reversed-phase columns with multiple attachment and lateral siloxane bridges.3 This improves the stability, but it is difficult to control the degree of polymerization induced by water3 and aggregation of particles is known to occur during silanization.4 By supplying the water adsorbed at the silica surface, these problems can be reduced.5,6 Several synthetic routes have been developed leading to a more stable silicon-carbon attachment bond. Among these is conversion of chlorinated silica with a Grignard or alkyllithium reagent.1,7-9 The Grignard reaction has, however, not been widely used for the preparation of chromatographic separation materials. The direct reaction of a Grignard reagent to the surface silanols of mesoporous silica at low pressure and high temperature10 has also been demonstrated. In hydrosilylation, introduced by Sandoval and Pesek for the preparation of chromatographic materials, the catalytic addition of a terminal vinyl group to a silica particle modified with a surface hydride was investigated11 and the concept further developed in a series of articles.12-14 Long alkyl chains have an ability to protect silica bonded phases to some extent against hydrolysis,15 and phases entrapping lower amounts of water in the bonded layer are more stable toward (3) Kirkland, J. J.; Glajch, J. L.; Farlee, R. D. Anal. Chem. 1989, 61, 2-11. (4) Ohno, K.; Morinaga, T.; Koh, K.; Tsujii, Y.; Fukuda, T. Macromolecules 2005, 38, 2137-2142. (5) Wirth, M. J.; Fatunmbi, H. O. Anal. Chem. 1992, 64, 2783-2786. (6) Wirth, M. J.; Fatunmbi, H. O. Anal. Chem. 1993, 65, 822-826. (7) Pesek, J. J.; Swedberg, S. A. J. J. Chromatogr. 1986, 361, 83-92. (8) Pesek, J. J.; Guiochon, G. J. Chromatogr. 1987, 395, 1-7. (9) Sunseri, J. D.; Gedris, T. E.; Stiegman, A. E.; Dorsey, J. G. Langmuir 2003, 19, 8608-8610. (10) Lim, J. E.; Shim, C. B.; Kim, J. M.; Lee, B. Y.; Yie, J. E. Angew. Chem., Int. Ed. 2004, 43, 3839-3842. (11) Sandoval, J. E.; Pesek, J. J. Anal. Chem. 1989, 61, 2067-2075. (12) Sandoval, J. E.; Pesek, J. J. Anal. Chem. 1991, 63, 2634-2641. (13) Chu, C. H.; Jonsson, E.; Auvinen, M.; Pesek, J. J.; Sandoval, J. E. Anal. Chem. 1993, 65, 808-816. (14) Montes, M. C.; van Amen, C.; Pesek, J. J.; Sandoval, J. E. J. Chromatogr., A 1994, 688, 31-45. (15) Poole, C. F. The Essence of Chromatography, 1st ed.; Elsevier: Amsterdam, 2003; p 286. 10.1021/ac0602874 CCC: $33.50

© 2006 American Chemical Society Published on Web 09/09/2006

hydrolytic attack.16 We are currently interested in phases for hydrophilic interaction chromatography (HILIC) with grafted polymeric chains. In HILIC, which was proposed by Alpert17 and is a subtechnique of normal-phase chromatography, the stationary phase should be highly hydrophilic. Alpert proposed a retention mechanism driven by partitioning between the bulk eluent and a water-enriched solvent layer on the stationary phase. The surface layer is consequently designed to maximize the water-retaining properties of the stationary phase, hereby also increasing its susceptibility to hydrolysis. Anchoring the polymer brushes by a hydrolytically stable bond is thus more important when making stationary phases for HILIC and other highly polar stationary phases. A large and increasing fraction of HILIC separations today is practiced on naked porous silica and is therefore encumbered with the inherent drawbacks of silica gel as a support in solidphase chromatography.15 Apart from the succinimide-modified silica phases pioneered by Alpert, only a handful of bonded phases have been developed specifically for use in HILIC, and with HILIC rapidly gaining interest in postgenomic research, new phases with different selectivity are needed. Low surface coverage and uneven film thickness can be overcome by using “grafting from” techniques, where an initiator moiety is immobilized on the substrate and polymerization is initiated from the surface. Surface-bound initiators can be prepared either by in situ synthesis on the surface18-20or by forming a selfassembled monolayer of initiator on the substrate.21-23 These methods produce polymer brushes with high polydispersity. Due to the fast polymerization rate, polymer chains growing inside pores will be starved for monomer at a relatively early stage since the major part of the monomer will be incorporated by the rapidly growing polymer chains at, or close to, the particle perimeter, eventually leading to complete blocking of the pore system.24 A relatively slow and controlled polymerization should make the polymerization rate less dependent on the diffusion path, provided the reaction rate is sufficiently impeded for the diffusion to compensate for local variations in monomer concentration. It should also result in a more uniform film thickness and decrease the risk of blocked pore orifices. One such controlled polymerization technique that has attracted a lot of attention in the past few years is atom-transfer radical polymerization (ATRP), pioneered by Matyjaszewski.25 In ATRP, the polymerization is achieved by homolytic cleavage of a halide atom from the growing terminal of the polymer chain. This cleavage is mediated by a transition metal complex (often CuBr complexed by two 2,2′-bipyridine molecules). After abstraction of the halide atom, the resulting Cu(II) complex acts as a deactivator and slows the polymerization rate. A more detailed look at the ATRP mechanism is beyond the scope of this article, but the (16) Hetem, M. J. J.; de Haan, J. W.; Claessens, H. A.; van de Ven, L. J. M.; Cramers, C. A.; Kinkel, J. N. J. Anal. Chem. 1990, 62, 2288-2296. (17) Alpert, A. J. J. J. Chromatogr. 1990, 499, 177-196. (18) Carlier, E.; Guyot, A.; Revillon, A. React. Polym. 1992, 16, 115-124. (19) Tsubokawa, N.; Ishida, H. J. Polym. Sci., Part A: Polym. Chem. 1992, 30, 2241-2246. (20) Frey, N.; Laible, R.; Hamann, K. Angew. Makromol. Chem. 1973, 34, 81109. (21) Prucker, O.; Ruhe, J. Langmuir 1998, 14, 6893-6898. (22) Prucker, O.; Ruhe, J. Macromolecules 1998, 31, 592-601. (23) Prucker, O.; Ruhe, J. Macromolecules 1998, 31, 602-613. (24) Revillon, A.; Leroux. D. React. Funct. Polym. 1995, 26, 105-118. (25) Wang, J. S.; Matyjaszewski. K. J. Am. Chem. Soc. 1995, 117, 5614-5615.

subject has been extensively reviewed.26-28 ATRP has several attractive features and was soon explored as an alternative chemistry for surface-initiated polymerization on silica and oxidized silicon.25,26 This is mostly achieved by first attaching a moiety containing an ATRP initiator by silanol condensation techniques similar to those previously used for making bonded-phase materials,29-35 and then the polymerization is carried out from this surface-tethered initiator. While this approach affords the advantages of controlled polymerization, it still suffers from the pH instability of conventional silica-based separation materials because the initial bond is a silicon-carbon ether. Other attachment procedures have also been utilized, based on thiol functionalization,36 on hydrosilylation,37 and by alternating polyelectrolyte layers.38 Further, since most ATRP initiators used also contain an ester linkage as part of the initiator moiety, the pH stability will be even lower. A novel way of initiating ATRP is directly from an oxidized and hydrated silicon surface, where the silanol groups have been replaced with chlorine atoms.39,40 This chlorine atom is highly reactive and cleaves from the silicone easily using the standard ATRP catalysts, producing a radical amenable to monomer incorporation. This initiation procedure has a number of benefits compared with the silane coupling approach. Preparation of chlorinated silica is simple and fast, and silica-initiated ATRP also forms a direct covalent bond between the silica substrate and the carbon polymer chain (a Si-C bond). In this work, we have employed a surface-initiated ATRP technique forming an initial Si-C bond for anchoring polymeric chains to porous silica particles. Using porous or fused silica as grafting substrate should make the chlorine even more reactive than chlorinated silicon or glass, and hence, there is an even greater need for totally anhydrous conditions, but it should also increase the initiation rate. EXPERIMENTAL SECTION Kromasil porous silica 5 and 10 µm, 200 Å from Eka Chemicals (Bohus, Sweden) was used as grafting substrates for all experi(26) Matyjaszewski, K.; Xia, J. Chem. Rev. 2001, 101, 2921-2990. (27) Singleton, D. A.; Nowlan III, D. T.; Jahed, N.; Matyjaszewski, K. Macromolecules 2003, 36, 8609-8616. (28) Pyun, J.; Kowalewski, T.; Matyjaszewski, K. Macromol. Rapid Commun. 2003, 24, 1043-1059. (29) Huang, X.; Wirth, M. Anal. Chem. 1997, 69, 4577-4580. (30) Ejaz, M.; Yamamoto, S.; Ohno, K.; Tsujii, Y.; Fukuda, T. Macromolecules 1998, 31, 5934-5936. (31) Chen, X.; Randall, D. P.; Perruchot, C.; Watts, J. F.; Patten, T. E.; von Werne, T.; Armes, S. P. J. Colloid Interface Sci. 2003, 257, 56-64. (32) Ejaz, M.; Tsujii, Y.; Fukuda, T. Polymer 2001, 42, 6811-6815. (33) Matyjaszewski, K.; Miller, P. J.; Shukla, N.; Immaraporn, B.; Gelman, A.; Luokala, B. B.; Siclovan, T. M.; Kickelbick, G.; Vallant, T.; Hoffmann, H.; Pakula, T. Macromolecules 1999, 32, 8716-8724. (34) Perruchot, C.; Khan, M. A.; Kamitsi, A.; Armes, S. P.; von Werne, T.; Patten, T. E. Langmuir 2001, 17, 4479-4481. (35) Granville, A. M.; Brittain, W. J. Macromol. Rapid Commun. 2004, 25, 12981302. (36) El Harrak, A.; Carrot, G.; Oberdisse, J.; Eychenne-Baron, C.; Boue, F. Macromolecules 2004, 37, 6376-6384. (37) Xu, D.; Yu, W. H.; Kang, E. T.; Neoh, K. G. J. Colloid Interface Sci. 2004, 279, 78-87. (38) Balachandra, A. M.; Baker, G. L.; Bruening, M. L. J. Membr. Sci. 2003, 227, 1-14. (39) Xu, F. J.; Cai, Q. J.; Kang, E. T.; Neoh, K. G. Macromolecules 2005, 38, 1051-1054. (40) Xu, F. J.; Cai, Q. J.; Kang, E. T.; Neoh, K. G. Langmuir 2005, 21, 32213225.

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Figure 1. XPS spectra of the chlorinated fused-silica surface and the same surface grafted with GMA. The peak at 200 eV is from Cl 2p, indicating covalently bound chlorine.

ments. Fused-silica capillaries (1 mm i.d. by 1.3 mm o.d.) were from Polymicro Technologies (Phoenix, AZ). The zwitterionic monomer 3-[N,N-dimethyl-N-(methacryloyloxyethyl)ammonium] propanesulfonate (SPE) was from Raschig (Ludwigshafen, Germany; used as received). 2,3-Epoxypropyl methacrylate (glycidyl methacrylate; 97%; deinhibited by passing through basic Al2O3) and butyl methacrylate (99%; distilled prior to use) were from Fluka (Buchs, Switzerland). Methanol (HPLC grade), thionyl chloride, copper (63 µm; >230 mesh), EDTA disodium salt (Titriplex III, p.a.), and potassium bromide (Uvosol) were from Merck (Darmstadt, Germany). The 2,2′-bipyridine, copper bromide, and methyl acrylate (99%; distilled prior to use) were from Aldrich (Deisenhofen, Germany). Dimethylformamide (Spectranal; DMF) and trichloromethane were from Riedel-de-Hae¨n (Seelze, Germany). Styrene (99%) was purchased from Acros (Geel, Belgium) and was distilled prior to use. Tetrahydrofuran was from JT Baker (Deventeer, The Netherlands), and acetonitrile was from Fisher Scientific (Loughborough, UK). Solvents used in preparation of the chlorinated silica were dried over 3-Å molecular sieves (VWR Scientific, Spånga, Sweden). Water was prepared in Ultra-Q equipment (Millipore, Bedford, MA) and had a resistivity of >15 MΩ cm-1. Preparation of the Silica Macroinitiator. The chlorination of silica was done according to the literature.20,41 An aliquot (2.05 g) of porous silica particles was refluxed in a mixture of 90 mL of (41) Jiang, W.; Irgum, K. Anal. Chem. 2002, 74, 4682-4687.

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thionyl chloride and 100 mL of trichloromethane under continuous stirring for 24 h. The chlorinated silica was collected by suction filtering on a sintered glass filter and briefly washed, first with trichloromethane and then with acetone, to remove most of the excess thionyl chloride. The silica was thereafter dried under vacuum, transferred to a glovebox, and kept under a nitrogen atmosphere. The original construction of the glovebox (Kebo, Stockholm, Sweden) was extensively modified with an airlock, and a moisture absorber consisting of a fan circulating the nitrogen through a tube with containing silica gel was situated inside the box. The particles were stored in a desiccator until used for grafting of a number of different monomers: styrene, methyl acrylate, butyl methacrylate, glycidyl methacrylate, and the zwitterionic monomer SPE. Grafting of GMA. Elemental copper (10 mg), 111 mg of 2,2′bipyridine, 75 mg of cuprous bromide, and 2.02 g of chlorinated silica were added to 40 mL of dimethyl formamide in a fluorinated ethylene propylene copolymer bottle with a tight fitting cap. At this point the solution turned dark brown, and 20 mL of GMA was added. After mixing all the chemicals, the bottle was tightly closed and removed from the glovebox for sonication (10 min) and then polymerization. Polymerization was carried out with the bottle attached to a custom-made rotor tumbling it end over end at 15 rpm for 1 h in a circulating air oven (Binder Fed series 720), maintained at 70 °C. The reaction was terminated by letting air into the bottle several times with intermittent shaking until the

Figure 2. Resolved C 1s XPS spectra of a GMA grafted silica surface.

solution turned bluish green. The particles were then collected by centrifugation and resuspended in THF. Three such washing steps with THF gave a clear solution with a blue pellet of particles. Two additional washings with 0.1 M aqueous Na2EDTA solution yielded a white particle pellet that was washed once with water and two more times with THF. The particles were then packed into a stainless steel column (150 × 2.1 mm) at 400 bar using a Haskel (Burbank, CA) DSTV-122 pneumatic amplifier pump with methanol as packing liquid. The columns thus prepared were subject to chromatographic evaluation. Grafting of SPE. SPE (6.95 g) was dissolved in methanol (89.2 g) and degassed with helium. CuBr (194 mg), 2,2′-bipyridine (210 mg), and chlorinated silica (1.5 g) were then added, in that order. Polymerization was carried out in the manner described above at room temperature for 48 h. Grafting of Other Monomers. Elemental copper (64 mg), 101 mg of 2,2′-bipyridine, 100 mg of CuBr, and 2 g of chlorinated silica particles were mixed with 6 mL of DMF. Two milliliters of this mix was then added to 4 mL each of styrene, butyl methacrylate, and methyl acrylate in 7-mL Pyrex glass tubes with tight-fitting caps. The mixtures were sonicated for 10 min and then polymerized at 90 °C for 24 h in the manner described above. The same washing protocol was also followed with the exception that toluene was used instead of THF for the washing of the styrene grafted particles. X-ray Photoelectron Spectroscopy (XPS). XPS was used to evaluate the chlorination and polymerization steps. The spectra were recorded with a Kratos (Manchester, UK) Axis Ultra photoelectron spectrometer using a mono Al KR source. To facilitate the XPS experiment, we used 1-mm-i.d. fused-silica capillary in ∼9-cm-long pieces. First, the polyimide coating was removed by pyrolysis in air at 600 °C; next the capillaries were etched using 1 M NaOH at 120 °C for 3 h, flushed with deionized water and acetone, and dried in an oven for 1 h at 120 °C, as described previously.42 The prepared capillaries were then moved into a capped vial while still hot and transferred to the glovebox. Since it was not possible to reflux inside the glovebox, two chlorination mixtures were applied; one consisted of thionyl (42) Courtois, J.; Szumski, M.; Bystro ¨m, E.; Iwasiewicz, A.; Shchukarev, A.; Irgum, K. J. Sep. Sci. 2006, 29, 14-24.

chloride/trichloromethane 1:1 (v/v) and the other was THF containing 1% (w/w) thionyl chloride and 0.1% (w/w) DMF. The second chlorination mixture was used since it is known to work at room temperature.39 Two pieces of capillary were immersed in each of the chlorination mixtures for 4 h at room temperature. Next, one capillary from each mixture was saved for XPS, and the other was subject to grafting with GMA according to the procedure described above. Prior to the XPS experiments, all capillaries were gently crushed in the glovebox and the largest pieces were placed on an XPS sample holder. The holder was then transported to the spectrometer using a glovebag filled with nitrogen. All operations with the capillaries were performed in such a way that contact with the laboratory atmosphere was kept at an absolute minimum, to avoid hydrolysis of the Si-Cl bonds. IR Spectroscopy. Sample preparation for IR was done by first drying the grafted silica under vacuum for 24 h at 60 °C. The samples were then mixed with KBr (weight ratio 1:50-1:100) and ground until homogeneous. IR spectra were collected in diffuse reflectance mode on a Bruker Equinox 55 (Ettlingen, Germany) fitted with a DRA-2CI cell from Harrick Scientific (Pleasantville, NY). The background (pure KBr) and sample signals were collected for 128 scans each between 5200 and 400 cm-1. Chromatographic Evaluation. A Bischoff (Leonberg, Germany) Compact HPLC pump and a Lambda 1010 UV detector were used for chromatographic evaluations. Samples were injected using a Rheodyne (Rohnert Park, CA) model 7125-081 injector fitted with a 0.1-µL PEEK loop. Data were collected with a Clarity Chromatography Station for Windows 2.3.0.148 from Data Apex (Prague, Czech Republic). For the reversed-phase separations, a 50/50% (v/v) mixture of methanol and water was used to separate four alkyl parabens (methyl to butyl) under isocratic conditions. The HILIC separations were made using an eluent consisting of 80% acetonitrile and 20% aqueous 25 mM ammonium acetate (v/ v). Hydrolysis of the Epoxy Groups. The column with the GMA grafted silica was further modified by hydrolyzing the epoxy groups to diol using 0.1 M H2SO4 pumped at ∼0.2 mL/min through the column that was heated on a water bath at 60 °C for 4 h. Total loss of retention for the mixture of alkyl parabens indicated that the hydrolysis was successful. Surface Area Measurements. Surface areas were measured by nitrogen adsorption/desorbtion on a FlowSorb II 2300 from Micromeritics (Norcross, GA) using the short flow path and a sample of ∼0.1 g. The samples were first dried at 120 °C for at least 3 h before being weighed into the adsorption/desorbtion chamber where they were further dried at 120 °C. Pyrolysis. The amount of grafted polymer on the porous silica was determined by first drying and weighing samples of both the GMA grafted and the hydrolyzed GMA grafted silica particles. The samples were then subjected to pyrolysis in air at 650 °C. The difference in weight before and after pyrolysis should correspond to the amount of polymer contained in the sample. RESULTS AND DISCUSSION The chlorinated silica beads were produced in a one-step reaction with thionyl chloride (this compound is very reactive and great care should be taken in its handling). Due to the rapid hydrolysis reaction of chlorinated silica, verification by XPS that the chlorination procedure actually leads to silica-bound chlorine Analytical Chemistry, Vol. 78, No. 20, October 15, 2006

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Figure 3. IR spectra of grafted porous silica before modification and after grafting. The peaks at 1730 cm-1 are from the carbonyl stretching in the (meth-)acrylate polymers. The absence of the silanol peak at 3740 cm-1 in all the grafted silicas is a strong indication that the silanol groups have been converted and are occupied as attachment sites for the grafts.

atoms had to be performed on chlorinated silica capillaries. If porous particles had been used, hydrolysis of the Si-Cl on the particle surface could occur during the drying step, which takes quite some time. The existence of covalently bound chlorine atoms on the silica surface was confirmed by the Cl 2p signal at a binding energy of 200 eV (Figure 1); however, the chlorination by thionyl chloride/trichloromethane 1:1 (v/v) was not very efficient without refluxing, so grafting of the capillaries was performed on substrates chlorinated with 1% (w/w) thionyl chloride and 0.1% (w/ w) DMF in THF. XPS was then performed on the capillaries grafted with GMA in the way described for the silica particles. The XPS spectra in Figure 2 show an increased carbon signal consisting of C-H and C-O lines of about equal intensity and a -COO signal component. The overall shape of the C 1s lines are comparable with those published previously,39 and we conclude that the surface had been covered by GMA polymer, presumably by ATRP grafting. FT-IR spectra of particles grafted with glycidyl methacrylate, styrene, methyl acrylate, butyl methacrylate, and the zwitterionic monomer SPE are shown in Figure 3. All methacrylate grafted particles show a peak at 1730 cm-1, which is specific for the ester bond linking the methacrylate side chains to the vinyl backbone, verifying that grafting had occurred. In the IR spectra for the nonmodified silica, the peak at 3740 cm-1 is from surface silanol groups. This peak is essentially absent in the spectra from the grafted samples, indicating that most surface silanol groups had been converted, corroborating the presumed high efficiency of the refluxing chlorination step in trichloromethane. The silanol peak is not affected if no grafting is achieved, i.e., in experiments where the grafting failed (data not shown). Differential gravimetric analysis of the pyrolyzed particles grafted with GMA showed that the weight increase upon grafting was 31%. The RP chromatographic behavior of the polyGMA grafted silica was also consistent with a thick grafted polymer 7102 Analytical Chemistry, Vol. 78, No. 20, October 15, 2006

layer, and the specific surface area decreased from 165 m2/g for the chlorinated silica to 95 m2/g for the GMA grafted silica. Slightly more than half (55%) of this difference can be explained simply by the weight increase of the particles, while the rest is accounted for by polymerization inside the silica pore system. The GMA grafted silica showed a reversed-phase retention pattern, evident from the increasing retention with alkyl chain length for the p-hydroxybenzoic esters used as test probes (Figure 4). The column efficiency was not satisfactory when tested in reversedphase mode, and a very low flow rate was necessary to obtain a reasonable chromatogram. Attempts were not made to improve the RP chromatographic properties since we (a) were not interested in this mode of separation and (b) the focus of this material paper is not on the chromatographic separations that can eventually be achieved in these initial syntheses but on the possibilities of the new controlled grafting technique. In the HILIC chromatograms (Figure 5) obtained from the same column after hydrolytic ring opening of the oxirane groups of the GMA into 2,3-propanediol functionalities, the efficiency was substantially improved, compared to the separations in RP mode. The pyrolysis weight loss of the HILIC stationary phase was 32%. This confirms that the loss of reversed-phase retention after hydrolysis was not caused by loss of grafted polymer chains. The high amount of grafting was probably the result of the addition of Cu(0) to the polymerization mixture, but unfortunately no grafting was detectable on silica particles without Cu(0) added. The only reasonable explanation is that the copper(I) bromide used was slightly oxidized and that hence the deactivation rate was too high. Addition of Cu(0) in ATRP has been shown to increase polymerization rate,43 presumably by reducing the Cu(II) species. (43) Matyjaszewski, K.; Coca, S.; Gaynor, S. G.; Wei, M.; Woodworth, B. E. Macromolecules 1997, 30, 7348-7350.

Figure 4. Reversed-phase HPLC separation of methyl, ethyl, propyl, and butyl p-hydroxybenzoate. Eluent: 50/50 (v/v) methanol/water at a flow rate of 7.5 µL/min. Column: 150 × 2.1 mm ATRP grafted GMA silica.

It is interesting to note that the hydrolyzed GMA HILIC phase showed different selectivity and superior peak capacity compared to the unmodified silica column (Figure 5). This demonstrates that there is still room for improvements in stationary phases for hydrophilic interaction chromatography and that grafted phases are a viable route in that direction. When the chromatograms are compared, it should be noted that the column packing might have affected the column efficiencies since it was not optimized for each stationary phase packed. CONCLUSIONS The surfaces of particulate and capillary silica-based separation materials can be chlorinated by either refluxing for 24 h in a 1:1 mixture of thionyl chloride and trichloromethane or, better, by treatment with 1% thionyl chloride and 0.1% DMF in THF at room temperature. Porous silica particles as well as fused-silica capillaries treated in this way can be used as macroinitiators for ATRP of a number of monomers, including glycidyl methacrylate, styrene, methyl acrylate, and butyl methacrylate. We were also able to graft the zwitterionic monomer SPE under these conditions, but a very high polymerization rate hints that the reaction may proceed by a different mechanism. This route provides one among few synthetic pathways leading to a direct Si-C bond between the silica and the grafted interactive

Figure 5. HILIC mode separation of toluene, uracil, and cytosine. Eluent: 80% acetonitrile/20% 25 mM ammonium acetate (v/v). Flow rate: 0.15 mL/min. Column: 150 × 2.1 mm ATRP grafted GMA silica after hydrolysis of the epoxy groups. Dashed line is nonmodified silica. Solid line indicates hydrolyzed GMA grafted silica.

layer. The main drawback of silica-initiated ATRP is the chlorinated silica, which is very reactive and makes handling under strict anhydrous conditions essential. ACKNOWLEDGMENT Eka Chemicals is gratefully acknowledged for supplying porous silica. We also appreciate the help of Per Persson and John Loring in acquiring and interpreting the IR spectra. This work was supported by the Swedish Science Research Council and by the Swedish Foundation for Strategic Research. Financial support from Stiftelsen J.C. Kempes Stipendiefond (P.H.) is gratefully acknowledged. An anonymous reviewer is also acknowledged for pointing out some additional key references. Received for review February 15, 2006. Accepted May 18, 2006. AC0602874

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