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Langmuir 1996,11, 4445-4448
In Situ STM and AFM Study of Protoporphyrin and Iron(II1) and Zinc(I1) Protoporphyrins Adsorbed on Graphite in Aqueous Solutions N. J. Tao,* G. Cardenas, F. Cunha, and Z. Shi Department of Physics, Florida International University, Miami, Florida 33199 Received April 20, 1995. In Final Form: July 27, 1995@ Iron(II1)protoporphyrin(IX), zinc(I1)protoporphyrin(IX), and protoporphyrin(1X)have been studied on the graphite basal plane in aqueous solutions with both scanning tunneling (STM)and atomic force (AFM) microscopies. Real-time images directly show that the molecules adsorb onto the electrode and condense into monolayer films starting from small islands. In the monolayer films, all three molecules lie flat on the surface and form identical two-dimensional lattices with a = 13.4 i 0.2 A , b = 12.2 f 02 A, and y = 68 i 2" . The corresponding molecular packing density is 1.1 x 10-lo mol/cm2,which is in excellent agreement with the value determined from cyclic voltammograms. Although the three molecules are nearly identical in their geometrical structures, their internal structures revealed by STM are significantly different. After the monolayer is completed,iron(II1)protoporphyrin(IX)forms aggregates with a thickness of -8.5 A, while zinc(I1)protoporphyrin(1X) and protoporphyrin(M) do not.
Introduction Iron protoporphyrin(1X) (Fe-PP) and related metalloporphyrins are of great interest both because of their critical roles in many biological processes and their electrocatalytic properties for reducing 0 2 . l These molecules have been studied by a variety of electrochemical and spectroscopic methods on m e r c ~ r y ,g~~- a~p h i t e , and ~-~ glassy carbonlo electrodes. In spite of these studies, the fundamental question of how the molecules are arranged on electrode surfaces has not been answered unambiguously. Experimental evidence has shown that Fe(II1)-PP molecules tend to aggregate into dimers or polymers in aqueous solutions,2 but it is not clear if the dimers or polymers remain intact after adsorbing on electrodes. While the measured molecular packing density on both mercury and graphite electrodes indicates that the molecules lie flat on the electrode^,^-^ a recent electroreflectance study has suggested that the molecules do not lie flat on the graphite electrode.8 Scanning tunneling (STM) and atomic force (AFM) microscopes have the capability of directly determining the molecular packing structure of adsorbates in aqueous solutions.11J2 These techniques have been used to study the molecular packing structure of several organic mol-
* To whom correspondence should be addressed. E-mail:
[email protected]. Abstract publishedinAdvanceACSAbstracts, October 1,1995. (1)The Porphyrins; Dolphin, V. D., Ed.: Academic Press: New York, @
1979.Electrochemical and Spectrochemical Studies ofBiological Redox Components. Kadish, K. M., Ed.; American Chemical Society: Washington, DC, 1982. Collman,J. P.; Denisevich, P.; Konai,y.;Marrrocco, M.; Koval, C.; Anson, C. F. J. A m . Chem. SOC.,1980,102,6027.Zagal, J.; Bindra, P.; Yeager, E. J. Electrochem. SOC., 1980,127,1506.Liu, H. Y., Weaver, M. J., Wang, C.-B.; Chang, C. K. J. Electroanal. Chem. 1983. 145. 439. (BjBednarski, T. M.; Jordan, J. J . Am. Chem. SOC.1967,89, 1552. (3) Kolpin, C. F.; Swoffrod, H. S. Jr.Anal. Chem. 1978,50,916,920. (4) Rusling, J. F.; Brooks, M. Y . J. ElectroanaL Chem. 1984, 163, ~
277 _ . ..
(5)Brown, A. P.; Anson, F. C. Anal. Chem. 1977, 49, 1589. (6) Shigehara, K.; Anson, F. C. J. Chem. Phys. 1982, 86, 2777. (7) Jiang, R.; Dong, S. Electrochem Acta, 1990, 35, 1227. (8) Sagara, T.; Takagi, S.; Niki, K. J. Electroanal. Chem. 1993,349, 159. (9)Arifuku, F.; Mori, K.; Muratani, T.; Kurihara, H. Bull. Chem. SOC.Jpn., 1992, 65, 1491. (10)Bianco, P.;Haladjian, J.;Draoui, K. J. Electroanal. Chem. 1990, 279, 305. (11)Liu, H. Y.; Fan, F. R. F.; Lin, C. W.; Bard, A. J. Am. Chem. SOC. 1986,108,3838.Sonnenfeld, R.; Hansma, P. K. Science 1986,232,211.
ecules adsorbed on electrodes in aqueous solutions with controlled electrode potential.13-15 Under ambient conditions, STM has been used to image porphyrin-based molecules on a gold substrate.16 In this paper, we report a combined STM and AFM study of Fe(II1)-PPon graphite adsorbed both from aqueous solution and from aqueous ethanol solution. We have also studied the adsorption of zinc(I1) protoporphyrin(1X) (Zn-PP) and protoporphyrin(1x1(PP)under similar conditions. The three molecules are nearly identical in the geometrical structure, but they are different in their electronic and chemical properties. So a comparison of the three molecules is of interest also in terms of understanding STM images of organic molecules.
Experiments The STM and AFM experiments were carried out on a Nanoscope I11system (DigitalInstruments). The STMtips were etched electrochemicallyfrom 0.25 mm diameter Pto.&o.z wires which were then coated with Apiezon wax. The AFM tips were commercially available Si3N4 tips supplied by Digital Instruments. Highly oriented pyrolytic graphite (Advanced Ceramic Co.) was used as the electrode on which the molecules are adsorbed. In each experiment, the electrode was covered with solution immediately after cleavage to reduce contamination.A Pt wire was used as a counterelectrode,and an Ag wire was used as a quasi-referenceelectrode which was calibrated against an SCE electrode. The voltammograms were measured on a model ADRF5 potentiostat from Pine Instruments Co. Fe-PP from Kodak and Zn-PP and PP from Aldrich were dissolved in 0.1 M NazB407 with pH adjusted to 10 by using NaOH. The concentrations of Fe-PP and Zn-PP were varied from 0.02 mM to 0.1 mM. A saturated PP solution was used because of its low solubility. The solutionswere prepared from a bioresearchgrade Nanopure water system (Barnstead) fed with campus distilled water. The STM experiments were performed both in air and in a Nz chamber,but the images were found to be essentially the same in both cases. (12)Drake, B.; Prater, C. B.; Weisenhorn, A. L.; Gould, S. A. C.; Albrecht, T. R.; Quate, C. F.; Cannel, D. S.; Hansma, H. G.; Hansma, P. K. Science 1989,243, 1586. Manne, S.; Hansma, P. K.; Massie, J.; Elings, V. B.; Gewirth, A. A. Science 1991,251, 183. (13) Srinivasan, R.; Murphy, J. C.; FainChtein, R. J. J . Electroanal. Chem. 1991,312, 293; Srinivasan, R.; Murphy, J. C.; FainChtein, R. J. Ultramicroscopy 1992, 42, 453. (14) Tao, N. J.; DeRose, J. A.; Lindsay, S. M. J . Phys. Chem. 1994, 98, 7422. (15) Tao, N. J.; Shi, Z. J . Phys. Chem. 1994, 98, 1464. (16) Luttrull, D. K.; Graham, J.; DeRose, J. A,; Gust, D.; Moore, T. A.; Lindsay, S. M. Langmuir, 1992,8, 765.
0743-746319512411-4445$09.00/0 0 1995 American Chemical Society
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Figure 1. Formation of a Fe-PP film on graphite from a 0.1M Na2B407 solution as monitored by AF'M. Before introducing Fe-PP into the cell, the image shows the bare graphite surface (A). 5 min after introducingFe-PP into the cell, a monolayer of Fe-PP has formgd (B). 10 min later, aggregates with a thickness of -8.5 A began to form on the monolayer (C).The images were obtained with an operation force of a few nanonewtons.
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Tao et al.
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Results Adsorption. We started the experiment by imaging a bare graphite electrode in 0.1 M Na2B407 solution with AFM or STM. After surveying a large area of the bare graphite surface, we introduced a drop of Fe-PP solution into the solution cell while the tip was still scanning. We then monitored the adsorption of Fe-PP onto the surface in real time. Figure 1A is an AFM image obtained before introducing Fe-PP into the cell, which shows a nearly atomically flat area of the bare graphite surface. Highresolution images of the area revealed the graphite lattice which was used as a standard for calibrating the lattice constants and for determining the relative orientation of the monolayers. A few minutes after introducing a drop of 0.1 mM Fe-PP into the cell, the molecules began to adsorb onto the surface and nucleated into many small islands. These islands grew gradually and coalesced into a monolayer film in a few minutes (Figure 1B). Waiting for longer time, the molecules continued to adsorb ontp the monolayer and form islands with a height of -8.5 A (Figure IC). These islands grew in size and population, but they did not condense into a uniform compact layer such as a monolayer; these will be referred to as aggregates. Rinsing the aggregate-coveredelectrode with 0.1 M Na2B407 did not remove the aggregates from the surface, which indicates a strong binding between the aggregates and the monolayer. It is well-known that Fe(II1)-PP can be reversibly reduced to Fe(I1)-PP on graphite, but it is not clear if the molecules both in the monolayer and in the aggregates participate in the electron transfer reaction. In order to answer this question, we have measured the voltammograms before and after the formation of the aggregates and found no significant difference in either the shape or the height of the peaks in the voltammograms. This indicates that only the molecules in the monolayer take part in the electron transfer reaction. For comparison,we have cari-ied out a similar study on Zn-PP and PP. The two molecules adsorbed onto the electrode and condensed into monolayer films in the same fashion as Fe-PP, but they do not appear to form aggregates after the monolayers are completed. Molecular Packing Structures. After studying the adsorption, we investigated the molecular arrangement in the monolayers with high-resolution STM and AFM. The AFM images are typically poorer in revealing the individual molecules than the STM images, so our discussions in this section will be based primarily on the STM images. Figure 2A is an STM image that shows ordered packing of the molecules in the monolayer of FePP. In addition to the ordered array, the image shows bright blobs due to the aggregatesdiscussed above. Higher magnificationSTM images (Figure 2B) show more clearly
Figure 2. STM images of a Fe-PP monolayer which show ordered packing of the molecules in the monolayer.Bright blobs pointed to by arrows in A are due to aggregates. The images were obtained with a tunneling current of 200 PA and bias of 100 mV. High-frequency noise in B has been removed.
Figure 3. Proposedmolecular packing structure in a monolayer of Fe-PP based on the STM images. A unit cell of the Fe-PP lattice is outlinedby solid lines, and the lattice directionsof the underlying graphite substrate are indicated by dashed lines.
the ordered packing of the molecules. From the STM images, the molecular packing density has been determined to be 1.1 x moVcm2, which is in excellent agreement with the value obtained from the cyclic volt ammo gram^.^ The lattice constaqts determined from the STM images are a = 13.4 f 0.2 A, b = 12.2 f 02 A, and y = 68 f 2" . The molecular lattice is rotated with respect to the substrate lattice by 20" as determined by comparing the Fe-PP image with the substrate image obtained before introducing Fe-PP into the cell. The molecular lattice is virtually identical in both Na2B407 and phosphate supporting electrolytes and independent of the concentration from 0.02 M to the saturated concentration. On the basis of these images, a model for the molecular packing of Fe-PP on the electrode is shown in Figure 3. In the model, the molecules lie flat on the surface and pack closely into a two-dimensional lattice. Using the bond lengths and angles from the X-ray crystallography data,17and the van der Waals radii from (17) Perutz, M. F.; Muirhead, H.; Cox,J. M.; Goaman, L. C. G.Nuture 1968,219,131.
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Figure 4. High-resolutionSTM images of Fe(II1)-PP(A), Zn(11)-PP(B), and PP (C)in their monolayers. The insets at the upper-right corners of the images are ball and stick models of the corresponding molecules. High-frequency noise has been removed from these images.
Figure 6. STM image of a line defect (twin dislocation)in the PP monolayer.
been observed in other organic monolayers grown from many small islands, and they have been shown by in situ AFM to be due to coalescence between neighboring i~1ands.l~So the line defects in the monolayers of the present system likely have the same origin.
Discussions Figure 5. Simultaneous STM image of PP molecules and the underlying graphite substrate lattice. The image was obtained with a tunneling current of 1nA and bias of 100 mV.
ref 18,theomodelproduces the following lattice constants: a = 14.2 A, b = 12.4 A, and y = 70°, which are in good agreement with the measured data. Since the propionic acid groups are actually bent out of the pyrrol-plane by -43",adjacent molecules appear to slightly overlap with each other when viewed from the top (Figure 3). We have also studied the molecular packing structure of the Zn-PP and PP monolayers and found that they pack into essentially the same structure as the Fe-PP monolayer. However, the internal structures of the three molecules revealed by STM are significantly different as shown in parts A-C of Figure 4. The differences are due to the differences in their electronic states. So STM can be used to distinguish structurally similar but electronically different molecules. In a recent study of guanine and xanthine, we also found a large difference in their STM images in spite of a remarkable similarity in their structure.l9 Most STM images were obtained with a tunneling current of less than -250 PA and a bias voltage of about 100 mV. With larger tunneling currents or smaller bias voltages, the Fe-PP monolayer and the underlying graphite lattice may be simultaneously imaged as shown in Figure 5. This kind of image provides direct information about the orientation of the Fe-PP lattice relative to the substrate lattice, which agrees with the result described above. The images also confirm that the molecules lie flat on the electrode. At very large tunneling currents or very small bias voltages, only the graphite lattice is visible. The monolayers are not perfect. They typically consist of many line defects (Figure 6). Line defects have also (18)Bondi, A.J. Phys. Chem. 1964,68,441. (19)Tao, N.J.;Shi, Z. J. Phys. Chem. 1994,98,7422. Tao, N.J.;Shi, Z.Surf Sci. Lett. 1994,341,L149.
The observation that the molecules lie flat on the graphite basal plane contradicts the conclusion of the recent UV-visible reflectance spectroscopy.8 This apparent contradictionis not due to different concentrations or buffer solutions used in the the reflectance and the present STM studies. We have prepared the sample using the same precedure as in ref 8 and imaged the sample in 0.1 M phosphate buffer, and we obtained similar images. One possible explanation is that reflectance spectroscopy measures the molecules both in the monolayer and in the aggregates, and molecules in the aggregates do not lie flat on the surface and thus are responsible for the measured polarization in the reflectance spectroscopy. Fe(II1)-PP was believed to form dimers in aqueous solutions. On mercury electrodes, it has been proposed that Fe(II1)-PP,in the form of dimers, is reduced to Fe(11)-PP via a two-electron-transfer reaction.2 However, the present STM study did not observe dimers of Fe(II1)PP in the monolayer on the graphite basal plane. A previous work found that the formation of dimers is prohibited in aqueous ethanol solution^.^ We have compared the Fe-PP monolayers adsorbed from 0.1 mM Fe-PP in 0.1 M NaB704 with that from 0.1 mM Fe-PP in 70% 0.1 M NaB704 30% ethanol and but found no significant difference in the molecular packing structure. This comparison further indicates that the molecules in the monolayer do not exist as dimers. We believe that the Fe(II1)-PPdimers are broken into monomers to pack into the ordered array when adsorbed on the graphite basal plane from aqueous solutions.
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Summary We have studied the adsorption and the molecular packing structures of iron(II1) protoporphyrin(M), zinc(11)protoporphyrin(M),and protoporphyrin(M) on graphite adsorbed from aqueous and from aqueous ethanol solutions with AFM and STM. The molecules adsorb on the electrodesurfacesand condense into monolayer films. The molecular-resolution images show that all three
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molecules pack into similar two-dimensional arrays in which the molecules lie flat on the electrode. The lattice constants are: a = 13.4 f 0.2A, b = 12.2 f 02A and y = 68 i 2" , leading to a molecular packing density of 1.1 x mol/cm2 which is in excellent agreement with the value determined from the Cyclic volta"ograms. The lattice of the is rotated the substrate lattice by -20" as determined by comparing the images of the molecular lattice with that of the substrate lattice. High-resolution STM images reveal the internal structures of the three molecules which are significantly
Tao et al. different from each other, indicating the difference in the electronic states ofthe molecules can be revealed by STM.
Acknowledgment. We thank Professors J. T. Landrum and J. M. Quirke for helpful discussions and for providing samples. This work was supported by Research Corporation (#C3608)and by the donors ofthe Petroleum Research Fund (#28163-GB7)administered by the American Chemical Society. LA950316Y
