J. Phys. Chem. C 2008, 112, 3741-3746
3741
In Situ Raman Spectroscopy of Redox Species Confined in Self-Assembled Molecular Films Nicola´ s G. Tognalli* and Alejandro Fainstein Centro Ato´ mico Bariloche and Instituto Balseiro, Comisio´ n Nacional de Energı´a Ato´ mica, 8400 S. C. de Bariloche, Rı´o Negro, Argentina
Carolina Vericat, Marı´a E. Vela, and Roberto C. Salvarezza Instituto de InVestigaciones Fisicoquı´micas Teo´ ricas y Aplicadas (INIFTA), UniVersidad Nacional de La Plata - CONICET, Sucursal 4 Casilla de Correo 16 (1900) La Plata, Argentina ReceiVed: August 3, 2007; In Final Form: December 13, 2007
In situ Raman spectroscopy and electrochemical experiments are used to study different methylene blue (MB) species immobilized in molecular films (sulfur, propanethiol, dodecanethiol, mercaptopropionic acid, and mercaptoundecanoic acid monolayers), self-assembled on SERS-active Au surfaces in aqueous electrolyte under potential control. The presence of three different MB species along reduction-oxidation cycles is inferred, irrespective of the nature of the molecular film. Residual Raman intensity in the spectroscopically silent leuco methylene blue (LMB) potential range demonstrate a dissimilar degree of connectivity for electron transfer between MB molecules and substrate depending on the spacer. In addition, the intensity of the Raman signal depends on the monolayer chemistry where hydrophobic and/or electrostatic interactions tend to stabilize the MB species against desorption from the molecular film driven by concentration gradients.
Introduction There is a great interest in the development of sensitive and specific chemical and biological systems due to their potential application in sensor, biosensor, and biocatalytic devices. In this context, the interaction between organic and inorganic material surfaces is a frontier topic. The development of these devices needs a precise control of the organic and inorganic interfaces in aqueous environments. Self-assembled monolayers of thiols have shown to be promising to link functional redox active molecules to metal surfaces.1 In fact, different types of active molecules have been supported on thiol-covered metallic substrates through covalent, electrostatic, or hydrophobic interactions. The study of the nanoscale properties of this type of interfaces (active molecule/thiol/metal) immersed in aqueous environments requires the use of an extremely sensitive technique like surface-enhanced Raman spectroscopy (SERS). SERS employs different types of nanostructured substrates to enhance the Raman signal produced by adsorbed and immobilized species,2-4 which is otherwise too weak to be detected with conventional Raman spectroscopy. Electrochemically roughened Au foils are one kind of these active substrates.5 In SERS, the effective Raman cross-section can be increased by many orders of magnitude. Therefore, this technique combines the ultrasensitive detection limits with the detailed structural information content of Raman spectroscopy. In this work we present a detailed in situ Raman and electrochemical study of methylene blue (MB) immobilized in 0.3-2 nm thick molecular self-assembled monolayers (SAMs) built of thiols or sulfur on electrochemical roughened Au foils. MB is an interesting molecule for biomimetic systems because of its similar redox behavior to NAD and FAD enzymes. * To whom correspondence should be addressed. E-mail: tognalli@ cab.cnea.gov.ar.
In previous studies we have explored these nanoscale systems by using voltammetric experiments6,7 and ex situ Raman spectroscopy.8 The voltammetric studies of MB immobilized on Au with sulfur (S) and with methyl- and carboxylateterminated thiols6,7 have concluded that the electron transfer to the substrate is much more efficient for S (measured charges are a factor of 10-20 larger than for the thiol SAMs), and that presumably for the thiol SAMs the charge transfer occurs through defects. On the basis of these studies, it was proposed that, for carboxylate-terminated thiols, MB molecules are mainly at the outer plane of the SAMs, “trapped” by electrostatic interactions. On the other hand, also on the basis of the electrochemical behavior of the system, some penetration of the MB molecules into the SAMs was suggested for methylterminated SAMs. However, voltammetric cycles provide a rather indirect measure of the amount of immobilized MB, because adsorbed MB molecules may not be close enough to the substrate for electron transfer to occur, thus remaining “silent” in such experiments. A direct measure of the presence of MB in the SAM may alternatively be obtained by Raman spectroscopy. In fact, ex situ Raman experiments performed on dry conditions confirmed that S is the most efficient SAM for MB immobilization, but demonstrated a MB surface concentration only 2-3 times larger than for the thiol-covered substrates.8 In these latter experiments, performed under SERS conditions, the decrease in the Raman intensity as a function of distance for rough Au electrodes was used to locate the average position of the MB species with respect to the Au substrate. These results confirmed that significant amounts of cationic MB species are able to diffuse into methyl-terminated thiols, but they are stopped at the outer plane of the SAM by negatively charged carboxylate groups of short hydrocarbon chains, like MPA. In the case of carboxylated thiols with longer hydrocarbon chains, such as MUA, an intermediate behavior was found, i.e., some MB
10.1021/jp076239k CCC: $40.75 © 2008 American Chemical Society Published on Web 02/16/2008
3742 J. Phys. Chem. C, Vol. 112, No. 10, 2008 molecules were driven between the thiol chains by lipophilic interaction while others were attached to the carboxylate groups by electrostatic forces. The comparison with the electrochemical experiments, however, is not straightforward. First, while the latter are done under wet conditions, the Raman experiments were taken on dry substrates immediately after the MB immobilization. Second, as stated above, while Raman scattering detects all the MB molecules present in the assembled films, voltammetric cycles are only sensitive to those molecules that are able to exchange charge with the substrate. To clarify these issues, we have performed a detailed combined in situ Raman and electrochemical study of MB immobilized on Au through S and different thiol SAMs. The detection of the Raman signal while the MB molecules change (or do not change) their electronic state along the electrochemical cycle allows for a direct identification of molecules that are able (or not able) to exchange charge with the substrate. Experimental Section We have performed experiments with MB immobilized on SERS-active rough Au substrates, modified by SAMs of different spacers: sulfur (S), methyl-terminated thiols (propanethiol (PT) and dodecanethiol (DT)), and carboxylateterminated thiols (mercaptopropionic acid (MPA) and mercaptoundecanoic acid (MUA)). The pairs PT and MPA, and DT and MUA, have basically a similar structure and length and only differ in their terminal group (nonpolar in the case of methyl, and polar in the case of carboxylate). The SERS-active substrates were prepared by the electrochemical roughening procedure described in ref 5. Briefly, a thick hydrous Au oxide film is formed by anodization of Au at a relatively high applied potential followed by voltammetric electroreduction. Au foils (99.99% pure) were used, that were immersed in a 0.5 M H2SO4 solution and held for 10 min at 2.4 V (vs a Ag/AgCl reference electrode). Finally, the potential was scanned down to -0.6 V at a rate of 0.02 V/s. The surface of the resulting electrodes consists of Au nano/microparticles with sizes ranging from 20 to 200 nm. The surface area of the electrodes was calculated by measuring the AuO monolayer electroreduction charge (q), considering that one monolayer involves 0.42 mC/cm2.5 Concerning the SAMs assembly, the solutions were prepared by using analytical-grade chemicals and Milli-Q water. First, the Au substrates were immersed in 0.1 mM PT, DT, MPA, or MUA ethanolic solutions for 24 h to form the corresponding SAMs.1,9-12 In the case of S, the Au substrates were immersed in 3 mM Na2S aqueous solutions saturated with nitrogen for 15 min. Under these conditions, the rectangular planar S8 surface structures are spontaneously formed on Au electrodes, as described in ref 13. The thiol-covered substrates were rinsed with ethanol and then with water, while the S-covered substrates were only rinsed with water. In order to test the spacer density on the SERS active Au we have estimated the electrodesorption charge of the S and thiol molecules from the rough surfaces, finding the same charge densities as for these SAMs selfassembled on flat Au(111) surfaces (0.15 mC/cm2 for S and 0.08 mC/cm2 for thiols).13,14 Immobilization of MB species on the spacer-modified Au substrates was performed by immersion for 30 min in freshly prepared 0.1 mM MB + 0.1 M NaOH aqueous solutions.6,7 Samples were rinsed with water and dried with a N2 gas flow. In situ Raman experiments were made in a conventional threeelectrode electrochemical cell using the MB-modified Au as
Tognalli et al.
Figure 1. SERS spectra of MB adsorbed on a S spacer on Au as a function of time (which is proportional to the applied potential) measured using 20 mW of the 647.1 nm laser line with 10 s of acquisition time. Scan rate 0.01 V/s.
the working electrode, a large Pt mesh as a counter electrode, and a Ag/AgCl reference electrode. The voltammetric runs were made typically from 0 V to -0.75 V at 0.01 V/s in aqueous 0.1 M NaOH solution. The cathodic limit was selected in order to avoid desorption of the alkanethiol monolayers. In fact, previous results using these nanostructured Au electrodes showed an increased stability for reductive desorption in these types of surfaces compared with Au(111).15 Regarding the reversible changes in the stable range, it should be noted that in situ STM studies of thiols on preferred oriented Au(111) single crystals showed only fluctuations between the x3 × x3 and the c(4 × 2) superlattices.16 The Raman experiments were made in situ performed using a Jobin-Yvon T64000 triple spectrometer operating in subtractive mode and equipped with a liquid-N2-cooled charge-coupled device. The excitation was done with a 647.1 nm Ar-Kr ion laser and a 750 nm Ti-Sapph laser. Typical powers were around 10 mW, concentrated on either a ∼100 µm diameter circular spot or a 7 mm long and ∼100 µm wide line focus. These were chosen to reduce the photon-induced degradation of the samples. Under the worst situation (circular spot), the photobleaching was determined to be around 5% after 15 s of data acquisition. To avoid accumulating this effect, a fresh spot in the sample was used after taking each spectrum. Prior to this, a Raman map of the samples was performed with very low powers and acquisition time to exclude spots displaying Raman intensities very different from the average value. To check for repeatability, several complete series of measurements were performed for MB immobilized on SAMs assembled with the five different spacers. Results and Discussion As an illustration of the typical experiment performed within this investigation, we present in Figure 1 the SERS spectra taken with 647.1 nm excitation, for MB immobilized on a sulfur SAM. The spectra were acquired as a function of the applied potential during a cyclic voltammogram in a 0.1 M NaOH aqueous solution. The Raman experiment was performed during the first electrochemical scan to avoid MB migration to the solution. The potential was scanned from 0 to -0.75 V, and back to 0 V, so that MB evolves from its natural oxidized state, to being fully reduced and finally back again to its oxidized form. The scan rate was 0.01 V/s, so each Raman spectrum is the MB average response within 0.1 V. This choice of scan rate and Raman acquisition time was the best condition found as a compromise to avoid both MB migration to the solution and
In Situ Raman Spectroscopy of Redox Species
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Figure 2. Solid curve: first voltammetric cycle of MB/S/Au interface, scan rate 0.01 V/s. Points: normalized intensity of the five major peaks labeled in Figure 1 as a function of the electrochemical potential, namely: a C-N-C skeleton bending (448 cm-1, labeled with a square), another C-N-C skeleton bending (500 cm-1, circle), a C-N symmetric stretching (1395 cm-1, up triangle), a C-N asymmetric stretching (1433 cm-1, down triangle), and a C-C ring stretching (1620 cm-1, rhomb), respectively. The dotted curve is a guide to the eye.
Figure 3. Normalized intensity of five major peaks of the MB SERS spectra for four different spacers PT (top left), DT (top right), MPA (bottom left), and MUA (bottom right) measured with the 647.1 nm laser line. The MB signals are labeled as in Figure 1: the C-N-C skeleton bending (448 cm-1, labeled with a square), the other C-N-C skeleton bending (500 cm-1, circle), the C-N symmetric stretching (1395 cm-1, up triangle), the C-N asymmetric stretching (1425 cm-1, down triangle), and the C-C ring stretching (1620 cm-1, rhomb). Dotted lines are a guide to the eye.
photobleaching, and getting Raman spectra with a good signalto-noise ratio. Spectra taken at fixed potentials and with longer integration times were also acquired, verifying that the scan averaging does not lead to undesirable artifacts or loss of significant information. Two main features can be highlighted from these spectra. First, the intensity of the peaks undergoes important changes as a function of the applied potential. And second, though the overall shape of the Raman spectra is conserved along the electrochemical experiment, some subtle changes (not evident in Figure 1) occur, suggesting the formation of different electrochemical-dependent MB species. These spectral changes in intensity and peak position will be discussed next. The intensity of the five major peaks, normalized to their initial value, is displayed in Figure 2 as a function of the electrochemical potential. These modes, indicated with the corresponding symbols in Figure 1, are the 448 cm-1 C-N-C skeleton bending (labeled with a square), the 500 cm-1 C-N-C skeleton bending (circle), the 1395 cm-1 C-N symmetric stretching (up triangle), the 1433 cm-1 C-N asymmetric stretching (down triangle), and the 1620 cm-1 C-C ring stretching (rhomb).17,18 Figure 2 also includes the electrochemical response (continuous curve) of the MB molecules adsorbed on a S monolayer on Au as a function of time (or applied potential, at a scan rate 0.01 V/s). It is clear from Figure 2 that all the displayed Raman peaks follow the same behavior as a function of the applied potential. First, an increase in the SERS intensity is observed with a maximum around -250 mV (25 s). This behavior could be caused by a modification of the electronic resonant Raman process when the molecule is reduced to the HMB• intermediate state. The weakness of the 470 cm-1 Raman signal compared to the peaks at 448 and 500 cm-1 in Figure 1 are strong evidence that the MB molecules adsorb mostly as dimers and not as monomers.8,19,20 This implies that the MB HOMO-LUMO optical absorption should be centered at 610 nm in the fully oxidized state. When the molecules are reduced, in a first stage this optical absorption edge red-shifts to 880 nm corresponding to the HMB• intermediate state.19,20 When the laser energy is above this latter transition, a strong resonant Raman condition may exist, giving rise to an additional enhancement mechanism (SERRS, for surface-enhanced reso-
nant Raman scattering).21 Note that the electronically resonant Raman process depends on whether the laser energy is above or below the optical absorption. In the former case, resonances occur at different excited vibronic states (Albrecht “A-term”) leading to a Raman intensity profile with maxima shifted above the absorption edge by repetitions of the vibration energy. Below the absorption edge, on the contrary, Raman efficiencies are typically smaller.21 As the magnitude of the applied potential is further decreased, the molecule is fully reduced to the LMB state at -750 mV (75 s), and the Raman intensity decreases to zero. For this molecular state, the optical absorption strongly up-shifts into the UV region.19 Under these circumstances the 647.1 nm laser line is so detuned from any electronic resonance Raman process that even with the SERS active Au substrate the Raman signal becomes very weak.21 Following the voltammogram in Figure 2, we observe again an increase in the intensity as we return to the HMB• potential at -250 mV (125 s), consistent with the SERRS mechanism described above. We note however that this second maximum displays only half the intensity observed in the reduction part of the cycle. This can be understood as due to a loss of adsorbed molecules to the electrolyte in the LMB state because of the smaller moleculesulfur interaction, as described by Campbell et al.22 This is in agreement with the Raman intensity observed at the end of the voltammetric cycle in the oxidized state E ) 0 mV (150 s), where only half of the initial intensity is recovered. In Figure 3 we present the SERS intensity vs electrochemical potential plots, similar to that shown in Figure 2 for S, but now corresponding to MB immobilized by the four thiolated Au substrates. The same peaks are analyzed as for S, except the one at 1433 cm-1 which, in the case of the thiol SAMs, we have observed that shifts to 1425 cm-1. All the curves display some common features. First, all the Raman peaks described in Figure 3 show essentially the same behavior along the electrochemical cycle displaying an intensity increase followed by a decrease as the MB molecules go through the HMB• and LMB states, respectively, similar to that depicted for S in Figure 2. Second, there is a relatively larger spread of values for MB immobilized by the thiol spacers than what was observed in the case of S. This could be implying that a larger distribution of molecular orientations (and thus Raman selection rules)
3744 J. Phys. Chem. C, Vol. 112, No. 10, 2008 affects the results when immobilization is through the larger molecular spacers. Further studies, however, are required to clarify this point. In addition, as we will discuss below, the intensity of the Raman signals is smaller for these thiol spacers when compared to S, thus leading to a reduced signal-to-noise ratio. On the other hand, some important differences between the panels of Figure 3 are also apparent. The more conspicuous being that, while for the carboxylate-terminated thiols (MPA and MUA) the Raman intensity goes almost to zero at the fully reduced state, for the methyl-terminated ones (PT and DT) this does not occur. The behavior for PT and DT is almost identical, showing a small peak related with the HMB• resonance tuning, followed by the depression characteristic of the LMB state. At this point, the lowest value reached by both curves is around 0.45. This means that a substantial amount of the MB molecules, which are present in the SAMs and thus lead to a Raman scattered signal, are not electrochemically connected to the Au substrate, and consequently their electronic resonance is not quenched by the reduction process. That is, around 45% of the MB molecules remain at their oxidized state (i.e., absorbing at 610 nm) even for the most negative potential, thus contributing significantly to the collected signal with laser excitation at 647 nm. This observation is consistent with the anomalously small electrochemical charge observed for these spacers when compared to the corresponding amount of Raman detected molecules, as was discussed in previous publications.6,8 Following the voltamperommetric cycle, the Raman intensity curves for PT and DT show a second peak again related with the HMB• state, acquiring finally in the oxidized MB+ state an intensity close to 0.7 of their initial value. This latter result suggests that some MB molecules desorb to the solution during the reduced LMB state, but with a smaller probability than for the S spacer. This is possibly due to van der Waals interactions between thiols and MB, and also because of the increased solubility of LMB molecules into hydrocarbon chains, diminishing the exposure to the NaOH solution as reported by Barner, B. J., et al.23 We now turn to the carboxylate-terminated thiols in Figure 3. MPA presents a shape similar to that of the methyl-terminated spacers but with a notable difference in the LMB (fully reduced) region where the intensity drops close to zero. This implies that for MPA, as for S, almost all the Raman detected molecules are electrically connected to the substrate. The same can be said for the longer carboxylate-terminated thiol, MUA. There are in addition two notable features in this latter case. First, the 1425 cm-1 C-N asymmetric stretching mode shows a distinctive behavior. We have not yet found any explanation for the almost insensitive dependence of this mode intensity with electrochemical potential. Second, there is a large increase in the intensity when the molecules are in the HMB• region, which is close to 2 during the reduction part of the cycle, but that reaches a comparatively larger value (close to 3.5) when the molecules are oxidized again. We will further address this latter peculiar point below. We have discussed in the previous paragraphs how the Raman intensity along the voltammetric cycles, through its dependence on the electronic structure of the molecule, can be used to probe the presence of MB in its different oxidation states. On the basis of this conclusion, the existence of residual Raman intensity at potentials corresponding to fully reduced states was presented as evidence of the existence of molecules that are not electronically connected to the substrate. This is consistent with previous investigations which show from voltamperometric cycles that the measured electrochemical charges are between 10 and 20 times greater when MB is immobilized on S as compared to
Tognalli et al.
Figure 4. Left panel: MB ∼ 1620 cm-1 SERS intensity ratio of samples with different spacers (S, PT, DT, MPA, and MUA) measured in NaOH 0.1 M solution and in air using the 647.1 nm laser line. Right panel: ∼1620 cm-1 SERS intensity of MPA sample measured in air, in H2O, and in NaOH using the 647.1 nm laser line. Each data point is the average of five measurements on every one of three samples built on the same way, and it is presented with its standard deviation.
those obtained on both methyl- or carboxilate-terminated thiols while Raman and SERS studies on dry substrates demonstrate the same trend but with MB concentrations for the S spacer that are only 2-3 times larger than those detected when using the thiol spacers.6,8 It is apparent from the present investigations that different quantities are measured in the two experiments. On the other hand, while performing the described experiments we noted that the initial intensity measured when immersing the samples for the electrochemical scans were different from those measured under dry conditions. That is, the lower electrochemical charges detected for the thiol spacers as compared to S could be due not only to the MB-substrate incomplete connection in the former but also by a different rate of desorption when the SAMs are immersed in the electrolyte. In order to test this hypothesis we performed a new set of experiments comparing the SERS intensities in air immediately after the molecular assembly with that detected in the same sample once immersed in the electrochemical solution at open circuit potential which is close to -0.2 V (in the MB+ region). In the left panel of Figure 4 we present the intensity ratio corresponding to the MB ∼ 1620 cm-1 mode between measurements performed on samples in a 0.1 M NaOH aqueous solution and in air. Each data point is the average of measurements performed over five different spot positions on three samples prepared at the same time and under the same conditions. The standard deviation is indicated with error bars. The results clearly demonstrate that MB+ desorbs to the solution with dissimilar proportion depending on the spacer. The SERS signal decreases to about 70% of the initial intensity for S, around 20-30% for PT and DT, to about 50% in MUA, and strikingly below 5% in MPA. An additional test to bring further light to these results is shown in the right panel of Figure 4, where we display the SERS intensity of the MB ∼ 1620 cm-1 peak for the particularly anomalous MPA spacer when the sample is immersed in air, in H2O, and in the NaOH 0.1 M aqueous solution (the latter corresponds to that used in the electrochemical studies). We observe that MB+ desorbs into H2O driven by diffusion forces which are important because MB+ is initially absent in the solution. However, the amount of MB+ transferred by diffusion to H2O is less than 30% of the initial value. On the other hand, the decrease in the MB+ Raman intensity when the sample is immersed in the NaOH solution reaches more than 95% of the initial value. We interpret that this drastic loss of MB+ follows
In Situ Raman Spectroscopy of Redox Species from two separate contributions, one related with the diffusion forces acting also in the case of H2O, and another, more important, caused by ion displacements. In other words, the Na+ ions in solution may be screening the charge exposed by the COO- terminal groups of MPA so those MB+ ions that were electrostatically attached to MPA are free to leave the SAM. This added mechanism explains why only a small fraction of the MB+ molecules remains on the MPA SAM, presumably inside defects or between the short carbon tails of the molecular layer. Within this model, we can explain most of the features demonstrated in Figure 4. First, MB+ adsorbed on S desorbs to the NaOH solution by the concentration gradient-induced diffusion, leaving 70% of the initial amount of molecules in the SAM. This same mechanism is more efficient for PT and DT SAMs for which only 20-30% of the molecules remain after immersion. This reduced desorption in S could be explained by the more effective molecular binding on S layers than on thiols, due to some contribution of the dimethylamino interaction with the sulfur layer.8,22 Concerning the carboxylate-terminated thiols, MPA and MUA, they also bind MB+ more effectively than their methyl-terminated counterparts (PT and DT) due to the electrostatic interactions with the polar end group. However, as discussed above, this attraction is screened and the molecules are replaced by the high concentration of Na+ ions in the 0.1 M NaOH solution, leading to the observed enormous MB+ mass loss to the electrolyte. As a result, less than 5% of the initial amount of MB+ remains assembled in MPA. The case of MUA, however, does not seem to agree with this general picture. Being a carboxylate-terminated thiol, this spacer should again demonstrate both the effects of diffusion and ion displacement. However, the Raman measurements in NaOH solution indicate more than 50% of the intensity measured in air. To clarify this issue, we recall that previously reported SERS measurements concluded from the average distance of the MB+ molecules to the substrate (that was significantly smaller than the spacer thickness) that MB+ is immobilized in MUA not only at the outer part by its polar attraction to the carboxylate end group but also that a significant penetration of molecules in between the MUA carbon chains has to occur.8 With this in mind, we can understand the difference observed with MPA in Figure 4, if we note that at the same time that the Na+ ions screen the charge of the COOterminal groups, they are repelling the remaining MB+ molecules, which are between the long hydrocarbon chains, down to the Au substrate. This process can be interpreted as an ion blockade of the MB+ molecules (which are initially between the carbon chains) produced by the Na+ ions, which inhibit their diffusion to the electrolyte and push them closer to the Au substrate, yielding an enhancement of the related Raman intensity due to the SERS strong dependence on molecule to metal distance. We note that, contributing positively to this latter mechanism, MB in its LMB form is more soluble in carbon chains.23 Besides providing an additional limit to the concentration-gradient-induced desorption, this effect could explain in a natural way the strikingly large and asymmetric maxima observed for MUA in Figure 3 as due to the added effect of the already discussed electronic Raman resonance, and the increased SERS amplification due to the movement of molecules toward the substrate in the LMB phase of the species. Besides the Raman intensity, the position of the peaks and their relative intensity in the Raman spectra can provide further information on the molecular changes occurring along an electrochemical cycle. We have performed a detailed study of MB with different spacers using both voltamperometric cycles
J. Phys. Chem. C, Vol. 112, No. 10, 2008 3745 and some fixed selected electrochemical potentials, the latter to better define the system condition and to allow for longer integrations times and thus better Raman signals. These experiments were performed using as laser excitation the 647.1 nm line of the Ar-Kr laser, and in some cases the 750 nm line of a Ti-Sapph laser to further resonantly enhance the spectra at intermediate potentials corresponding to the HMB• state of methylene blue. All the most intense Raman peaks follow essentially the same trend for the five studied spacers. In all cases, these changes are reversed when the potential returns to 0 V, and the molecules are again fully oxidized. The ∼1620 cm-1 C-C stretching shows a downshift to ∼1612 cm-1 following the potential from 0 to -0.75 V, while on the same scan the ∼1425 cm-1 C-N asymmetric stretching shows an upshift of about ∼5 cm-1 and the ∼1395 cm-1 C-N symmetric stretching shifts down by ∼8 cm-1. On the other hand, the C-N-C skeleton bending vibrations at ∼448 cm-1 and ∼500 cm-1 do not demonstrate any substantial shift. Besides these mode shifts, in the case of MB adsorbed on S, DT, and MUA when the molecule is reduced to its HMB• form, we observed a new peak around ∼1132 cm-1. The latter has been assigned by Nicolai et al. to a N-H bending involving the added H atom that bonds to the central N during the HMB• and LMB oxidation states.19 Presumably, this new H atom slightly modifies the central ring structure because of the positive charge delocalization,19 leading to the observed shifts of the 1620, 1425, and 1395 cm-1 peaks. Within this description, the lower energy bending vibrations would be less affected than the stretching ones by the molecular variations induced by the bonded H atom. On ending, we note that in some cases the shift of the ∼1620 cm-1 C-C ring stretching is accompanied by the appearance of a second peak at ∼1626 cm-1. At this point we should note that as we were using the 647 nm and the 750 nm laser lines for the experiments with all the spacers, the weak shifted signals we measured at -0.75 V are presumably related to MB molecules in a remaining HMB• state, probably due to oxygen traces in solution, and not to MB in its LMB state which is only absorbing and thus resonant in the UV. Conclusions We have used SERS spectroscopy to study the different methylene blue species formed in five different molecular films in aqueous electrolyte following electrochemical cycles. Evidence obtained from the intensity of the Raman signals, related to the electronic structure of the molecule, indicates that three different species are formed depending on the applied potential. On the other hand, the spectral data, which is related to the structure of the molecules, can only probe two of the oxidation states, MB and HMB•. The differences in the residual Raman intensity at the LMB potential region for the various spacers show an unequal degree of connectivity for electron-transfer processes between the MB molecule and the Au substrate. Furthermore, from the variation of the SERS-measured intensities under dry and wet conditions with different solutions, and for the diverse molecular spacers, we have demonstrated that the latter exhibit dissimilar degrees of MB loss to the solution. For NaOH aqueous solution, this desorption depends basically on two mechanisms: (i) the concentration-gradient-induced molecular diffusion, present in all the cases, and (ii) the ion displacement, acting only for the carboxylate-terminated thiols. We also found SERS evidence that a MB migration into the hydrocarbon chains exists for the MUA spacer when the sample is in NaOH solution. This could be due to an ion blockade and an electrostatic repulsion between the positive charge of the MB+ and Na+ ions.
3746 J. Phys. Chem. C, Vol. 112, No. 10, 2008 Acknowledgment. We acknowledge financial support from ANPCyT (PICT02-11111) and CONICET (PIP 6075). This work was done within the framework of the Interfacial, Supramolecular and Molecular Nanoscience and Nanotechnology Net and Nanoscience and Nanotechnology Net, Nanostructured Materials and Systems, Argentina. M.E.V. is a member of the research career of CIC. References and Notes (1) Wilbur, L.; Whitesides, G. M. Self-assembly and Self-assembled Monolayers in Microand Nanofabrication. In Nanotechnology; Timp, G., Ed.; Springer-Verlag: New York, 1999. Cunningham, A. J. Introduction to Bioanalytical Sensors; Wiley-Interscience: New York, 1998. (2) Kneipp, K.; Wang, Y.; Kneipp, H.; Perelman, L.; Itzkan, I.; Dasari, R. R.; Feld, M. S. Phys. ReV. Lett. 1997, 78, 1667-1670. (3) Maher, R. C.; Cohen, L. F.; Etchegoin, P. Chem. Phys. Lett. 2002, 352, 378-384. (4) Tognalli, N.; Fainstein, A.; Calvo, E.; Bonazzola, C.; Pietrasanta, L.; Campoy-Quiles, M.; Etchegoin, P. J. Chem. Phys. 2005, 123, 044707. (5) Salvarezza, R. C.; Arvia, A. J. A Modern Approach to Surface Roughness Applied to Electrochemical Systems. In Modern Aspects of Electrochemistry; Conway, B. E.; Bockris, J. O. M.; White, R. E., Eds.; Plenum Press: New York, 1996; Vol. 28, p 289. Alonso, C.; Salvarezza, R. C.; Vara, J. M.; Arvia, A. J.; Vazquez, L.; Bartolome´, A.; Baro´, A. M. J. Electrochem. Soc. 1990, 137, 2161-2165. (6) Benitez, G.; Vericat, C.; Tanco, S.; Remes Lenicov, F.; Castez, M. F.; Vela, M. E.; Salvarezza, R. C. Langmuir 2004, 20, 5030-5037. (7) Vericat, C.; Remes Lenicov, F.; Tanco, S.; Vela, M. E.; Salvarezza, R. C. J. Phys. Chem. B 2002, 106, 9114-9121.
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