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Probing the Reaction Pathways of DL-Proline on TiO2 (001) Single Crystal Surfaces Gary J. Fleming and Hicham Idriss* Department of Chemistry, University of Auckland, Private Bag 92019, Auckland, New Zealand Received February 26, 2004. In Final Form: May 25, 2004 The reaction of DL-Proline on O2-annealed (stoichiometric) and O-defected (sub-stoichiometric) TiO2 (001) single-crystal surfaces has been investigated. This is of significance in trying to understand the concept of how biomolecules interact with the surfaces of biomedical implants (molecular recognition). On an O2-annealed TiO2 surface, proline is found to largely adsorb then desorb intact at ≈350 K. DFT (B3LYP) calculations of proline bound to a Ti(OH)4 cluster suggest a binding through the carboxylate functional group rather than through the NH group of the ring. In contrast, proline reaction was considerably different on the O-defected surface. First, proline was further stabilized, evidenced by a shift of its desorption temperature (during temperature-programmed desorption) to ≈530 K. Along with proline desorption, two distinctive sets of reaction processes occurred at 530 and 630 K, respectively. The first pathway (R) at 530 K shows desorption of large amounts of m/e 55 (attributed to 1-azetine) and m/e 42 (attributed to ketene). At still higher temperature, 630 K, a pathway (β) dominated by the appearance of low masses, mainly m/e 28, 27, and 26, is seen. These masses are tentatively attributed to desorption of HCN, ethylene, and/or acetylene as they represent the logical further decomposition of the different fragments of proline.
1. Introduction
* Autho to whom correspondence should be addressed. Fax: 64 9 373-7422. Tel: 64 9 373 7599 ext. 88760. E-mail: h.idriss@ auckland.ac.nz.
Most studies under ultra-high vacuum (UHV) conditions have involved the investigation of amino acids, such as glycine or alanine, on a variety of metallic surfaces, such as Cu or Pt single crystals.6,7,11,12 Although these studies are fundamentally interesting, neither copper nor platinum are used as biomaterials; therefore, this gives little information on the reactivity and binding of these biomolecules on a biological implant. Few works have addressed the reactions of amino acids on TiO2 surfaces. Lausmaa and co-workers have studied the adsorption and coadsorption of water and glycine on a TiO2 thin film13 by thermal desorption spectroscopy (TDS). They have found that, at coverages greater than 1 monolayer, desorption of molecular glycine occurred from the surface at approximately 300 K. At coverages less than a monolayer, desorptions at approximately 450 and 600 K became noticeable. The authors have assigned this feature to the dissociation followed by recombination of glycine (450 K) and surface reaction products such as CO2 (600 K). Soria and co-workers have investigated the binding of glycine on a TiO2 (110) crystal surface using high-resolution photoelectron spectroscopy.14,15 They have found that the multilayers adsorb via a zwitterionic configuration due to the presence of three bands in the ultraviolet photoelectron spectrum at 27.6 (O 2s, carbonyl), 24.1(N 2s, NH3+), and 18.1 eV(C 2s, R-carbon). When the surface was bombarded with photons, the coverage of glycine was reduced to less than two multilayers. Most of the glycine has desorbed, and the remaining layer is formed by C, NHx (x ) 1, 2), and OH hydroxyl species, as well as the formation of Ti3+ surface species.
(1) Albrektsson, T.; Brånemark, P. I.; Hansson, H.; Kasemo, B.; Larsson, K.; Lundstro¨m, I.; McQueen, D.; Skalak, R. Ann. Biomed. Eng. 1983, 11, 1. (2) Long, M.; Rack, H. J. Biomaterials 1998, 19, 1621. (3) Jones, F. H. Surf. Sci. Rep. 2001, 42, 75. (4) Kasemo, B. Surf. Sci. 2002, 500, 656. (5) Castner, D.; Ratner, B. D. Surf. Sci. 2002, 500, 28. (6) Zhao, X.; Zhao. R. G.; Yang,W. S. Langmuir, 2000, 16, 9812. (7) Chen, Q.; Frankel, D. J.; Richardson, N. V. Surf. Sci. 2002, 497, 37. (8) Kokoli, M. E.; Biesalski, M. Surf. Sci. 2002, 500, 61. (9) Meng, M.; Stievano, L.; Lambert, J. F. Langmuir 2004, 20, 914.
(10) Langel, W.; Menken, L. Surf. Sci 2003, 538, 1. (11) Barlow, S. M.; Kitching, K. J.; Haq, S.; Richardson, N. V. Surf. Sci. 1998, 401, 322. (12) Chen, Q.; Frankel, D. J.; Richardson, N. V. Surf. Sci. 2002, 497, 37. (13) Lausmaa, J.; Lo¨fgren , P.; Kasemo, B. J. Biomed. Mater. Res. 1999, 44, 227. (14) Soria, E.; Roma´n, E.; Williams, E. M.; de Segovia, J. L. Surf. Sci. 1999, 433-435, 543. (15) Soria, E.; Roma´n, E.; Williams, E. M.; de Segovia, J. L. Surf. Sci. 2000, 451, 181.
The use of titanium metal implants in the aiding of healing fractures in teeth and bone is commonplace in modern medicine. The choice of titanium as an implant material is based on both its mechanical properties, such as young’s modulus, and on its relative chemical inertness.1,2 The titanium metal’s main function is to supply a surface for which biomolecules can adhere to and to aid in the natural healing of the fracture site present in the body.3 Once placed in the body’s aqueous environment, the implant undergoes an oxidation process where the formation of a thin oxide layer in the range of 10-100 nm thick occurs. This thin oxide layer is crucial since it prevents the Ti metal from reacting with the biological molecules. However, the nature of interaction of the biomolecule (such as a protein) with this thin titanium oxide surface at the molecular level will ultimately determine its conformation. If the conformation of the biomolecule is altered from its naturally occurring state, it may cause the body to undergo an autoimmune response and reject the implant.4,5 This process is known as molecular recognition. It is the goal of understanding these types of complex interactions of proteins, enzymes, and other biomolecules in biological systems which has led to the fundamental investigation of their building blocks, amino acids, on various polycrystalline and single-crystal surfaces.6-10
10.1021/la049492+ CCC: $27.50 © 2004 American Chemical Society Published on Web 07/30/2004
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2. Experimental Section
Figure 1. Reconstructed 011 surface of a TiO2(001) single crystal. Black circles represent Ti atoms, and white circles represent O atoms. The shortest Ti-Ti distance of 3.57 Å is too long to accommodate a carboxylate group in a bridging configuration.
There are two reported surface science studies of S-proline on a Cu (110) single-crystal conducted by the same group16,17 but none on TiO2. The use of reflectionabsorption infrared spectroscopy (RAIRS) technique has led to information of how the ring system is orientated at the surface through the isolation of vibrational band position for each individual CH2 and CH group in the pyrrolidine ring. Proline is bound to the surface in an anionic fashion via both the carboxylate function and the imino group present in the ring. The low-energy electron diffraction (LEED) pattern shows that proline adopts a (4 × 2) pattern at both high and low coverages due to the structural rigidity displayed by the adsorped molecule. Tempurature-programmed desorption (TPD) studies have shown proline to be stable up to 450 K, at which point it undergoes dehydrogenation followed by decomposition at 535 K, and the following, largely unidentified, masses were detected: m/e 2, 27, 28, 29, 40, 41, 43, 44, and 67. In this paper, we present the investigation of the surface chemistry and binding of DL-proline on a TiO2 (001) stoichiometric and substoichiometric single-crystal surface using TPD. Proline was chosen as it is a constituent of collagen I, which is a major high-tensile structural protein found in teeth, bone, and cartilage.18 It is this protein that is generally found at the bone/implant interface site. This makes this system interesting to study from a biomaterials point of view, as we can gain an understanding as to how bone itself will interact with the implants’ surface. The chemistry of a large number of organic compounds on the rutile TiO2 (001) single-crystal surfaces has been investigated previously and is presented in three recent review articles.19-21 The use of a single-crystal surface was chosen as, unlike polycrystalline surfaces, the coordination of both Ti and O are well-defined. The TiO2 (001) single-crystal surface has all Ti atoms 4-fold coordinated and all O atoms 2-fold coordinated. This structure is unstable, and upon annealing at 750 K, the thermodynamically stable {011}faceted structure is formed (Figure 1). The surface of the {011} facet contains all Ti atoms 5-fold coordinated to oxygen. The surface created by argon ion-bombardment does not show a LEED pattern and contains a large number of O defects, and as a result, some Ti ions are present in lower oxidation states than +4, with a distribution depending on the sputtering conditions.19,20 (16) Barlow, S. M.; Ravel, R. Surf. Sci. Rep. 2003, 50, 201 and references within. (17) Mateo Marti, E.; Barlow, S. M.; Haq, S.; Raval, R. Surf. Sci. 2001, 501, 191. (18) Metals as Biomaterials; Helsen, J. A., Breme, H. J., Eds.; Biomaterials Science and Engineering Series; Wiley & Sons: West Sussex, 1998. (19) Barteau, M. A. Chem. Rev. 1996, 96, 1413. (20) Idriss, H.; Barteau, M. A. Adv. Catal. 2000, 45, 261. (21) Diebold, U. Surf. Sci. Rep. 2003, 48, 53 and references within.
All TPD experiments were performed in a UHV chamber described previously.22 The chamber is equipped with a quadrupole mass spectrometer (QMS) (up to 300 amu) with a Pyrex shroud, an ion sputter gun, an x, y, r motion feed through manipulator, an ion gauge, and two dosing lines with a dosing needle positioned near the surface of the crystal face. The TiO2 (001) single crystal was mounted to a roughened 1 cm2 piece of Ta foil 0.25 cm thick via a ceramic glue (Ceramabond 571). The crystal was then suspended between two copper rods by two Ta wires (0.5 mm diameter), which were then spot-welded to the foil. The temperature of the crystal was monitored through a K-type thermocouple attached to the side of the crystal. The TiO2 (001) crystal surface was cleaned via several sputtering and annealing cycles. Sputtering was carried out using Ar+ (2 × 10-5 Torr, 25 mA emission current and a 4 kV beam voltage). Once sputtered, the crystal was flashed and then annealed in O2 (1 × 10-5 Torr, 10 min at 800 K) for several cycles. XP spectra were recorded in a separate UHV chamber pumped with an ion pump, titanium sublimation pump, and a sorption pump to a base pressure in the 10-10 Torr range. The main chamber is equipped with Perkin-Elmer, dual anode X-ray source (magnesium and aluminum, KR 1253.6 and 1486.6 eV, respectively), angle resolved double pass cylindrical mirror analyzer, an electron beam for Auger spectroscopy (Perkin-Elmer phi model 11-010), and a sputter system (Perkin-Elmer phi model 04-191). The DL-proline, contained in a glass-metal sealed bulb, was prepared by heating at 90 °C while pumping (≈10-2 Torr), for a period of 2 h. The bulb temperature was then raised to 150 °C to obtain enough vapor pressure to dose (approx 1 Torr). We have noted that 150 °C was a high enough temperature to obtain the required vapor pressure to dose and low enough to prevent the decomposition of proline. Other workers have indicated, using an IR/matrix isolation technique, that at 150 °C proline was not decomposed.23 TPD experiments were carried out on both stoichiometric and substoichiometric surfaces at various coverages by altering either the dosing pressure of proline or the exposure time. The surface exposure was from 90 to 0.6 L (1L ) 10-6 Torr s) at 300 Ksthe exposure was not corrected for the ionization efficiency for proline. Substoichiometric surfaces were prepared by sputtering the surface with Ar ions. Conditions used in sputtering the crystal were varied to look at the effect of surface reduction on proline adsorption. The sputtering conditions were Ar+ pressure ) 2 × 10-5 Torr, emission current ) 25 mA, and beam voltage ) 3 or 4 kV for different periods of time. Confirmation of proline (m/e 115) being introduced into the chamber was performed by monitoring its the major ions: m/e 71, 70 (major fragment), 69, 68, 67, 43, 42, 41, 29, and 28, using the QMS. The intensity of these masses was used to form the basis of a cracking pattern to help assign desorption peaks due to molecular proline. The following fragmentation patterns were also used during data analyses: hydrogen cyanide (m/e 27, 26), ethylene (m/e 28, 27, 26), ketene (m/e 42, 41, 28, 14), propene (m/e 42, 41, 40, 39, 38, 27), and 1H-pyrrole 2,3-dihydro (m/e 69, 68, 43, 41) and 2Hpyrrole (m/e 67, 66, 41, 39). Numerous studies have shown that, in the gas phase, proline (and in general amino acids) is in its neutral form.25,26 Upon interaction with ions (in the gas phase) the amino acid can be in a zwitterionic or nonzwitterionic form depending on the nature of interaction27-32 (more details are given in the Results and (22) Wilson, J. N.; Titheridge, D. J.; Idriss, H.; J. Vac. Sci. Technol. A 2000, 18, 1887. (23) Stepanian, S. G.; Reva, I. D.; Radchenko, E. D.; Adamowicz, L. J. Phys. Chem. A 2000, 105, 10664. (24) Hehre, W. J. A Guide to Molecular Mechanics and Quantum Chemical Calculations; Wave function Inc.: Irvine, CA, 2003. (25) Stepanian, S. G.; Reva, I. D.; Radchenko, E. D.; Adamowicz, L. J. Phys. Chem. A 2001, 105, 10664. (26) Czinki, E.; Csa´sza´r, A. G. Chem. Eur. J. 2003, 9, 1008. (27) Marino, T.; Russo, R.; Toscano, M. J. Phys. Chem. B 2003, 107, 2588. (28) Hoyau, S.; Ohanessian, G. J. Am. Chem. Soc. 1997, 119, 2016. (29) Jockusch, R. A.; Lemoff, A. S.; Williams, E. R. J. Am. Chem. Soc. 2001, 123, 12255. (30) Jockusch, R. A.; Price, W. D.; Williams, E. R. J. Phys. Chem. A 1999, 103, 9266.
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Figure 2. TPD spectrum of selected mass fragments of proline on stoichiometric {011} faces of the TiO2 (001) single crystal. Surface coverage at or near saturation; proline exposure > 10 L. Discussion section below). To aid in our TPD analysis, calculations involving the study of L-proline was conducted on a Ti(OH)4 cluster. Theoretical calculations of the different types of adsorption processes of l-proline to a Ti(OH)4 cluster were performed
using the Spartan 02 computational chemistry program version 1.02 with density functional theory (DFT) methods (B3LYP) with the 6-31G* basis set.24 A pseudopotential was used to aid the speed of the calculation. This type of calculation is not intended to conduct full computational studies on the surface but was mainly designed to help in the interpretation of the TPD data. The energy reported (E) is that of the difference in energy between the model, Ti(OH)4/L-proline (Emodel), and the sum of the energy of l-proline (EL-proline) and that of Ti(OH)4 (ETi(OH)4) after energy minimization, for each using the same procedure; E ) Emodel (EL-proline+ ETi(OH)4).
3. Results and Discussion The main peaks desorbing during proline TPD on the O2-annealed TiO2(001)-surface is shown in Figure 2. All masses were monitored (up to 100 amu) in separate runs (12 masses in each run); in addition to these masses, a few mass fragments above m/e 100, such as 115, 140, and 168, were also monitored to determine if any coupling products were being produced. An example of the possible coupling products that might be expected is given in Scheme 1. No coupling products were seen. The selected masses desorbing at ca. 340 K, shown in Figure 2, are all attributed to proline. This assignment is based on the comparison of the peak area ratios of masses 70:71, 70:69, 70:68, and 70:43 to that of a reference gas-phase mass spectrum of DL-proline.33 As can also be seen, some desorption of m/e 43 occurs above 500 K. Other masses (such as traces of m/e 41 and 39) were also seen to desorb at this temperature domain. These are due to some decomposition of proline on minor surface defects as will be shown later. (31) Hoyau, S.; Ohanessian, G. C. R. Acad. Sci. Paris 1998, 1, 795. (32) McGill, P.; Idriss, H. work in progress. (33) http://webbook.nist.gov.
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Figure 3. Calculated optimized structures for the adsorption of proline on a Ti(OH)4 cluster. Calculations were performed at the B3LYP/6-31G* level using a pseudopotential. Three types of adsorption were modeled (a) nondissociative adsorption to the titanium atom through the carboxylate group, (b) dissociative adsorption to the titanium atom through the carboxylate group, and (c) dissociative adsorption to the titanium atom through the pyrrolidine nitrogen.
Before further analysis of the reaction pathway was done, we have found it worthwhile to perform theoretical calculations to aid in the understanding of how proline is interacting with the TiO2 surface. Proline has mainly two possible binding sites for which it can adsorb to the surface of TiO2: (i) coordination of the proline to the titanium metal center through the carboxylate function or (ii) coordination through the nitrogen atom of the fivemembered pyrrolidine ring. From the resulting simulations, three possible structures were obtained for the adsorption of proline and their respective binding energies. These structures are displayed in Figure 3. The three modeled adsorption processes to titanium are (a) nondissociative adsorption of proline through the carboxylate functional, (b) dissociative adsorption through the carboxylate functional, and (c) adsorption through the dissociated pyrrolidine nitrogen. For the nondissociative adsorption through the OH group of the carboxylate functional (Figure 3a), the binding energy for proline to the Ti(OH)4 molecule was very weak, -57 kJ mol-1. The calculation of the geometries and binding energies for the two dissociative adsorption processes involved the transfer of the H from either the OH (Figure 3b) or NH group (Figure 3c) to one of the OH groups on Ti(OH)4, thus forming, in essence, a Ti(OH)3(H2O) complex. We have opted for OH groups instead of O groups to maintain charge neutrality and because the surface of TiO2 in an aqueous environment contains substantial amounts of terminal Ti-OH, terminal TiOH2+, and Ti-OH+-Ti groups.35 The binding energies calculated for proline bonding through the dissociation of N-H of the ring and of O-H of the carboxylic groups were -86 kJ mol-1 and -138 kJ mol-1, respectively. This suggests that the there is a stronger affinity for the proline bonding to the titanium through the dissociated carboxylate species. We have opted to disregard a binding of proline via both the N and the O atoms to the surface Ti on the basis of data from extensive gas-phase computational studies of amino acids with ions. In brief, only very small ions, such as Li+27 and Be2+32,34, could make the bonding with both O and N atoms of amino acids. Large ions were always attracted to the two O atoms of the carboxylate functional27,29,32. Moreover, the steric effect of the ring would most likely disfavor such an arrangement on the (34) Strittmatter, E. F.; Lemoff, A. S.; Williams, E. R. J. Phys. Chem. A 2000, 104, 9793. (35) Connor, P. A.; Dobson, K. D.; McQuillan, A. J. Langmuir 1999, 15, 2402.
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Scheme 1. Possible Coupling Products (m/e 140 and 160) that May Be Formed upon the Adsorption/Reaction of Proline onto the Stoichiometric {011}-Faceted Surface of the TiO2 (001) Single Crystal
surface. It is also worth indicating that, on TiO2, NH3 is weakly adsorbed to the surface when compared to carboxylic acids. For example, the heat of adsorption of ammonia calculated from TPD results was found to be equal to -83 kJ mol-1 on this same single crystal.36 Other workers have found a very similar heat of absorption for NH3 on a TiO2(110) single crystal.37 On the other hand, the dissociative heat of adsorption of acetic acid on this same crystal was found to be far higher (-250 kJ mol-1 38). Patthey and co-workers have performed XPS of biisonicotinic acid (2,2-bipyridine-4,4-dicarboxylic acid) on a rutile TiO2 (110) single crystal.39 Bi-isonicotinic acid has the same possible binding sites that proline has available, namely nitrogen-containing rings and carboxylate groups. Submonolayer studies have shown through the use of XPS of the O 1s peak that there are two peaks present at 535.0 and 536.6 eV. The peak at 536.6 eV is attributed to the adsorption of deprotonated oxygen atoms of the carboxylic group binding to titanium. Nitrogen binding was investigated using X-ray adsorption fine edge structure, but no π-orbital resonance was found at 398.5 eV, which is characteristic for a nitrogen ring bound to the surface. It is, thus, highly likely that, given the above results and our data, proline is adsorbed on the stoichiometric (O2-annealed) TiO2 surface through the carboxylic group. As stated previously, there are two distinctive desorption regions for proline on the oxidized surface with the highertemperature desorption region (above 500 K) attributed to proline undergoing further reactions on TiO2 surface. This tends to suggest that there are surface states which are available to facilitate the binding and reaction of proline itself. Figure 4a shows a Ti 2p region composed of (almost exclusively) Ti4+ cations, indicating that defects are in trace amounts. The data reported in Figure 3 was acquired on a surface similar to that shown in Figure 4a. Figure 4b shows a surface which has been deliberately sputtered with Ar+ ions causing the creation of Ti atoms in lower oxidation states than 4+. The reduced surface has been studied previously40-42 and has been reported to be behind a large number of reduction reactions, including (36) Wilson, J. N. Ph.D. Thesis, The University of Auckland, 2004. (37) Farfan-Arribas, E.; Madix, R. J. J. Phys. Chem. B, 2003, 107, 3225. (38) Titheridge, D. J.; Wilson, J. N.; Idriss, H. Res. Chem. Intermed. 2003, 29, 553. (39) Patthey, L.; Rensmo, H.; Persson, P.; Westermark, K.; Vayssieres, L.; Stashans, A.; Petersson, A.; Bru¨hwiler, P. A.; Siegbahn, H.; Lunell, S.; Mårtensson, N. J. Chem. Phys. 1999, 110, 5913. (40) Idriss, H.; Lusvardi, V. S.; Barteau, M. A. Surf. Sci. 1996, 348, 39. (41) Idriss, H.; Barteau, M. A. Catal. Lett. 1996, 40, 147. (42) Idriss, H.; Pierce, K. G.; Barteau, M. A. J. Am. Chem. Soc. 1994, 116, 3063.
the formation of formaldehyde from formic acid,40 butene from acetaldehyde,41 and stilbene from benzaldehyde.42 The overall stoichiometry of the surface and near-surface was found to be equal to 1.65 (computed form the corrected XPS O(1s) to Ti(2p) peak area ratios). To investigate the effect of surface defects on the trace desorption above 500 K shown in Figure 2, we have performed TPD experiments after sputtering the surface with Ar+ followed by annealing in O2 (1 × 10-6 Torr) at 750 K for different times. Figure 5 shows the desorption of m/e 70; for simplicity, the desorption of the reaction products will be presented in details in Figure 6. As can be seen on the reduced surface (TiO1.8), there is the presence of only one desorption region at ∼550 K. As the experiment is repeated on the oxygenannealed surfaces, we see the decrease in prominence of this feature and begin to see a peak appear at lower
Figure 4. XPS spectrum of O2-annealed (1 × 10-6, 750 K), ≈TiO2, and Ar+-sputtered, TiO1.65, Ti 2p peak of TiO2 (001) single-crystal surface. Surface and near-surface stoichiometries of the defected surface is calculated from XPS peak area ratios of the O(1s) to Ti(2p) with the assumption that that the O2-annealed surface is stoichiometric or very near to stoichiometric.
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Figure 5. The effect of O2 annealing on the position and intensity of proline desorption (m/e 70); proline exposure ≈15 L.
Figure 6. Peak desorption during TPD following adsorption of proline at 300 K on a TiO2(001) single crystal that has been Ar ion bombarded for 45 min prior to adsorption; proline exposure ≈15 L
temperatures. After repeat annealings, we have complete removal of the desorption peak at 550 K and the reappearance of the desorption peak at ∼350 K associated with the molecular desorption of proline from an oxidized surface. This shift of the desorption temperature of proline on the reduced surface to the oxidized surface can be attributed to the replacement of the oxygen sites on the crystal surface, thus, decreasing the number of stronger binding sites available for the proline to bind to.
Fleming and Idriss
From the mass tracking experiments performed on the reduced TiO2 surfaces, several key masses were found to be present and are shown in Figure 6. The surface was deliberately bombarded with Ar ions for 45 min to create large amounts of surface defects (with a computed stoichiometry ) TiO1.65). Proline appears to undergo two distinct surface reaction pathways, labeled R and β. The R pathway occurs at approximately 530 K with contributions from several masses, including 70, 69, 55, and 42. The desorption of m/e 55 and 42 is of particular interest as these mass fragments are not a characteristic fragments of proline, therefore, tending to suggest that their presence is a surface-related phenomenon. The shoulder of the R peak (clear on m/e 55) is indicative of a more complex desorption that might be coverage dependent. The β pathway that occurs approximately 100 K higher in temperature produces high amounts of low molecular masses: m/e 28, 27, and 26. The desorption temperature of the R peak is coverage dependentsit shifts to lower temperature with higher coverage. Figures 7 and 8 show the effects of increasing surface exposure to proline prior to the running of TPD. The exposure in langmuir indicated on the figure is approximate since the pumping speed is relatively slow for Proline. It is, however, clear that at low coverage one desorption peak is seen, while a shoulder becomes prominent with increasing exposure. Above 10 L, the main desorption peak has either shifted by 30 K to lower temperature or was overshadowed by another peak. This shift is not due to the presence of multilayer because of several reasons. First, multilayers should not be favored at such a high temperature. Second, m/e 55 also follows the same trend as m/e 70, and the former is not resulting from proline fragmentation. A similar observation is seen for m/e 42 (not shown). The appearance of the second peak can be attributed to either reconstruction of the adsorbed proline layer on the surface or the adsorption on several distinctly different sites. The last attribution is more realistic because the surface is amorphous and contains Ti ions in several oxidation states associated with its large amounts of oxygen defects (cluster defects). Inspection of the possible reaction products from proline and the different desorptions was conducted, and identification of the various components contributing to both the R and β pathways was made by comparing the intensities of the various peaks for each mass with those of reference mass spectra.33 On the basis of this information, the following appear as the most likely surface reactions. The desorption of m/e 42 is accompanied by a desorption of m/e 41 and 28 at the same temperature (these masses are characteristic of ketene). Ketene is very often observed from the decomposition of carboxylic acids and its formation is not surprising.22,38,43 The large desorption of m/e 55, the reaming fragment, re-enforce this assignment. It is also possible that a decomposition via 2Hpyrrole, 3,4-dihydro (m/e 69) occurs in association with a decarboxylation of proline (CO/CO2 + H2O/H2). However, the 2H-pyrrole, 3,4-dihydro does not give a mass of 55. Another stable product that may also occur is the 2Hpyrrole (m/e 67) that has been seen to decompose to HCN and propyne by other workers.44 We did see only traces of m/e 67 (the parent ion of this molecule), and in addition, 2H-pyrrole does not give m/e 55.33 We have attributed m/e 55 to 1-azetine. There are no reported mass spectrometer data for 1-azetine. However, McAllister has reported (43) Kim, K. S.; Barteau, M. A. J. Catal. 1990, 125, 353. (44) Somers, K. R. F.; Kryachko, E. S.; Ceulemans, A. J. Phys. Chem. A 2003, 107, 5427.
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Figure 7. Effect of increasing proline exposure on m/e 70 and 55 desorption peaks. Scheme 2. Different Possibilities for Surface Reaction during Proline TPD onto an Ar+-Sputtered TiO2(001) Single-Crystal Surfacea
Figure 8. Computed (noncorrected) TPD peak areas of the main desorption fragments m/e 70, 69, 55, 41, 39, and 29 as a function of the surface exposure to DL-proline.
the presence of protonated 1-azetine through the use of FID/MS.45 This compound is highly unstable, yet several theoretical and vibrational studies have been conducted.46-49 Moreover, it has been reported that 1-azetine can isomerize to 2-azabutadiene (also unstable upon heating).50 2-Azabutadiene would then decompose into ethylene and HCN. The desorption of m/e 28, 27, and 26 at higher temperature (above 600 K) is most likely linked to this further decomposition. From the above data, the most probable end products resulting from proline decomposition at low surface exposure are ketene (at 550 K) followed by ethylene, HCN, and water (above 600 K). Some desorption of CO2 was also seen, and because of the contribution of ethylene and ketene into CO, the desorption of the latter cannot be excluded. In Table 1, the relevant masses observed during TPD experiments on reduced surfaces with their measured (and corrected peak) areas are displayed. Corrected peak areas (45) McAllister, T. Aust. J. Chem. 1984, 37, 511. (46) Amatatsu, Y.; Hamada, Y.; Tsuboi, M. J. Mol. Spectrosc. 1987, 123, 267. (47) Guillemin, J.-C.; Denis, J.-M.; Lasne, M.-C.; Ripoll, J.-L. Tetrahedron 1998, 44, 4447. (48) Sugie, M.; Takeo, H.; Matsumura, C. J. Am. Chem. Soc. 1989, 111, 906. (49) Bacharch, S. M.; Liu, M. J. Org. Chem. 1992, 57, 209. (50) Guillemin, J.-C.; Denis, J.-M.; Lablache-Combier, A. J. Am. Chem. Soc. 1981, 103, 468.
a The route to 1-azetine (framed) is the most plausible one based on the mass spectrometer signal.
Scheme 3. Isomerization of 1-Azetine into 2-Azabutadiene Followed by Decomposition to Ethylene and HCN
are presented to show the relative contribution of each particular compound. Correction factors are computed following the method described previously by Ko and coworkers.51 A corrected peak area was unable to be given for m/e 55 as there is no recorded mass spectrum for this 1-azetine. In Table 1, we present a breakdown of the mass fragments detected from the TPD experiment shown in (51) Ko, E.; Benziger, J.; Madix, R. J. J. Catal. 1989, 62, 264.
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Table 1. Observed Masses and Measured and Corrected Peak Areas for the Adsorption of Proline on a Reduced TiO2(001) Single-Crystal Surface mass fragment (m/e)
contribution from specific product
70 69 55
proline proline 1-azetine/ 2-azabutadiene ketene ethylene CO ethylene HCN ethylene HCN acetylene
42 28 27 26
peak area (Torr)
Scheme 4. The Creation of Surface Oxygen Defects Results in a Strong Adsorption for Prolinea
corrected peak area (Torr)a
6.0 × 10-8 4.3 × 10-8 1.0 × 10-7
5.5 × 10-7
4.3 × 10-8 4.3 × 10-8
7.7 × 10-8 4.3 × 10-8 c
3.0 × 10-8
3.0 × 10-8 c
4.4 × 10-8
4.4 × 10-8 c
1.0 × 10-7 b
a Corrected peak area found by multiplying the peak area by a correction factor. The correction factor takes into account the ionization efficiency, the gain of the multiplier, and the transmission through the quadrupole and is conducted relative to m/e 28, as outlined in ref 50. b Correction factor not available. c Peak areas are not corrected because of possible overlap (contribution) of several products.
Figure 6. We have given the decomposition product that contributes to a particular mass fragment and the amount of the product present. Figure 4 shows that, on the defected surface, proline is very stable since it desorbs (and reacts) above 500 K. The strong adsorption is due to the creation of surface oxygen defects. It is not unconceivable to consider that one of the oxygens of the carboxylic function (in the case of a free proline, it is most likely the hydroxyl oxygen, but in the case of proline in a peptide bond, it would be the oxygen of the carbonyl) is incorporated into the lattice upon adsorption as in the scheme below. This mode of adsorption would considerably change the structure of proline. Although making a generalization from this observation to a real protein adsorption in vivo cannot be done without simplification, a change in the structure of a protein containing a large number of prolines, such as collagen, will become inevitable.
a Although two modes for oxygen incorporation from proline may occur (O of the hydroxyl group and O of the carbonyl group), that of the carbonyl group is preferentially drawn here since, in a peptide chain, it is the only one available.
4. Conclusions Molecular biorecognition is important for the successful design of metal implants (such as Ti implants). A number of proteins in close proximity to the Ti implant (such as in collagen I, a structural protein found in teeth, bone and cartilage) contain a large percentage of the amino acid proline. The reaction of proline on the surface of the stoichiometric and reduced TiO2(001) single-crystal surfaces was conducted in order to see into the effect of surface defects on its reaction pathways. On a stoichiometric surface (O2 annealed), proline does not seem to decompose, and the only probable reaction is the deprotonation of the carboxylic group. In contrast, the creation of surface defects has a major effect on the adsorption/reaction of proline. Proline appears to be strongly adsorbed on the defected surface (a shift of the desorption temperature from 350 K on the stoichiometric surface to 530-550 K on the oxygen-defected surface). The strong adsorption energy, triggered by the presence of O-defects, results in the decomposition of proline into HCN, ketene, and ethylene/ acetylene. The decomposition to HCN and ethylene is most likely to occur via the formation of a four-membered ring (1-azetine) that may isomerize to 2-azabutadiene before further decomposition. Semiquantitative analysis of desorption products indicates that the decomposition pathway has contributed ∼1/3 of the total surface coverage. LA049492+
