Observation of Au–S Interface of L-Cysteine on Gold Surface

We studied the metal-molecular interface of Au–S bonds at the L-cysteine modified gold surface using X-ray photoelectron spectroscopy (XPS) and near-edge x-ray absorption fine structure (NEXAFS). Two kinds of Lcysteine layers, i.e., multi-layer and two-layers, were prepared on gold substrate. The formation of unique chemical bond was found at the sulfur-gold interface. The sulfur 1s core-level energy of two-layered L-cysteine on gold is higher by 8.2 eV than that of multi-layered sample. The large chemical shift was also observed in S K-edge NEXAFS spectra. Such remarkably large chemical shift in the case of two-layers has never been reported for any sulfur compounds. The direction of the S 1s energy shift for Au-S bond is opposite to that for the normal metal sulfides. This means that the electrons are donated from the sulfur atom to the gold substrate, and most likely the sulfur atoms are positively charged at likely [δ] states. We conclude that the strong Au-S bonds that are recently used for the immobilization of molecules are due to the observed specific Au–S bond. [DOI: 10.1380/ejssnt.2009.110]


I. INTRODUCTION
The chemistry of sulfur (S)-containing molecules on surfaces has attracted much attention due to the potential application for organic thin film devices such as self assembled monolayers (SAMs) [1].It has been well reported that the S-containing molecules like alkane thiols (R-SH) are adsorbed on well-defined metals [2,3], such as Au(111) and such adsorption often leads to well ordering of molecules at surfaces.On the other hand, few studies have been performed on multi-functional SAM molecules like L-cysteine[HSCH 2 CH(NH 2 )COOH], which is one of the amino acids containing S-H, amino (NH 2 ) and carboxylic (COOH) groups, although it is a candidate material of highly ordered layers due to lateral interaction.For instance, thiols(R-SH) on Au surface are the prototypical self-assembled monolayers (SAMs) system in which Au-S covalent bonding anchors the molecules to the surface.As for bio-molecules, proteins or artificial peptides have been shown to bind to gold surfaces through sulfur atom [4,5].L-cysteine containing a sulfur atom in the molecule itself is a unique amino acid forming a chemical bond with Au surface through interaction of SH group with Au atoms [6,7].It has been reported that L-cysteine molecule dimerizes with L-cysteine pairs on Au (110) surface [8].Moreover, several different adlayer structures of L-cysteine on the Au (111) were reported by different groups [7][8][9].They reported that L-cysteine formed (4 × √ 7)R19 structure on Au (111) surface.Recently, structures of surface and interfaces have been extensively investigated by Scanning Probe Microscopy (SPM), and the method has been applied to the observation on biomolecules on metal surfaces.Although the spatial resolution of SPM is atomic level, SPM allows mainly the observation surface morphology, thus there remain difficulties in clarifying the bonding state between adsorbed molecules and substrate directly with SPM [12][13][14].Corelevel spectroscopies such as XPS and NEXAFS are ef- * Corresponding author: honda.mitsunori@jaea.go.jp fective to research metal-molecular interface.NEXAFS is a suitable method for the study of chemical state of Au-S bond, since it provides information about the electronic structures of both core levels and valence unoccupied states that are localized at the S atom.Especially, NEXAFS at the S K-edge has advantages [15][16][17][18] over that at the S L-edge because the core-to-valence resonance peak directly probes the valence unoccupied 3p* states due to the dipole selection rule.
In this study, we report on the S-Au interface bond state between L-cysteine and Au surface using S K-edge NEXAFS and S 1s XPS analysis.The origin of the specific chemical bond is discussed on the basis of the observed energies of S 1s core levels and valence unoccupied states for both two-layer and multi-layer films.

II. EXPERIMENTAL
All experiments were carried out at the BL 27A soft Xray station in the Photon Factory of the High Energy Accelerator Research Organization (KEK-PF), Japan.The double crystals of InSb (111) were used as a monochromator.The energy resolution of the monochromator was 1.1 eV at 2.5 keV (sulfur K-edge).This beam line has the end station consisting of two ultra high vacuum systems (UHV system), i.e., main analyzing chamber and sample preparation chamber.The base pressure of both chambers was in the order of 10 −7 Pa.The main analyzing chamber consists of a hemispherical electron energy analyzer (VSW Co. Class-100) for XPS measurements and manipulator.The preparation chamber was equipped with Knudsen-cell (K-cell) evaporator, quartz crystal thickness monitor and sample transfer system.
NEXAFS spectra were measured by a total electron yield (TEY) obtained by the sample drain current.The incident angle of the X-ray beam was 90 • from the surface.The photon energy was calibrated by using Au 4f peak in the XPS spectra.
L-cysteine was purchased from TOKYO KASEI KO-GYO Co., Ltd (purity: 99.9%).The thin films were grown by vacuum deposition method using K-cell evaporator at 130 • C. We used the polycrystal Au clean sample in all experiment.The surface of Au was rinsed with acetone before introducing into the vacuum chamber and then the surface was sputtered by 1 keV Ar + ions.The energy of Ar + ions is quite low (under 1keV), so the ion beam causes no beam damages on the surface.We prepared two kinds of films on this polycrystal Au substrate.One is multilayer and the other is two-layer.The thickness of the films was determined by the XPS measurements.We will present the details of the estimation of the film thickness in the result and discussion part.In the XPS spectra, the intensities of the C 1s, N 1s, O 1s and S 1s peaks normalized by the photoionization cross sections of the respective orbitals are in good agreement with the atomic composition of L-cysteine.This confirms that no contamination such as hydrocarbon or water exists and the molecules are not decomposed during the evaporation.

III. RESULTS AND DISCUSSIONS
Before showing the NEXAFS results, we first describe how we evaluated the thickness d of the film that we prepared using the vacuum evaporation method.As expected, the I(S 1s)/I(Au 4f ) ratio increases with an increase in sample thickness.For homogeneous thin films covering the gold substrate, the intensity of the S 1s photoelectron, I(S 1s), is expressed as where K is a constant that depends on the detection efficiency and the x-ray flux, σ(S 1s) is the photoionization cross section of S 1s for 2472 eV photons, λ L-cys (S 1s) is the inelastic mean free path (IMFP) of the S 1s photoelectron in L-cysteine, and n(S) is the atomic concentration of sulfur in the L-cysteine samples [19].For gold, the Au 4f photoelectrons I(Au 4f ) are calculated as where all of the parameters are the same as those in Eq. ( 1).For δ, the reported values were used [19].Using the TTP-2M equation [20], the values λ L-cys (S 1s), λ Au (Au 4f ) and λ L-cys (Au 4f ) were estimated to be 8.05, 11.44 and 18.5 nm, respectively.I(S 1s)/I(Au 4f ) ratio in the XPS spectrum is 0.0251, so we obtain d = 1.25 nm.
Considering that the height of one monolayer is 0.591 nm [21], the film thickness of this sample is estimated to be two molecular layers.(Hereafter we call the twolayers).In addition, another sample is also estimated to be about twenty molecular layers using same method.
(Hereafter we call the multi-layers).S K-edge NEXAFS spectra for multi-layered and two-layered L-cysteine are shown in Figs.1(a) and (b), respectively.In multi-layered sample, a sharp peak is observed at 2475.0 eV (marked A).It was reported that this peak originates from the resonant excitation from S 1s to valence unoccupied σ * orbitals that are localized at the S-C bond [22].(Hereafter we call this peak as S 1s → σ * (S-C)).On the other hand, in two-layered sample, a sharp peak is located at 2484.0 eV (marked B), which is higher by 9.0 eV than that for multi-layered L-cysteine.For multilayered sample, it is considered that NEXAFS spectrum at the S K-edge represents the molecular L-cysteine, because most of the sulfur atoms do not interact with gold even if the sulfur atoms in L-cysteine at the top surface layer chemically interact with gold.The high energy peak observed in the NEXAFS spectrum for two-layered sample (peak B) suggests that strong chemical bonds are formed between sulfur atoms and gold.We can know the core-level shift by S 1s XPS measurement.
Figure 2 shows the S 1s XPS spectra for L-cysteine on Au surface.The spectra (a) and (b) represent multilayered and two-layered samples, respectively.For multilayered sample, only one main peak (marked A) is observed at 2472.2 eV.It is presumed that this peak corresponds to the S 1s of molecular L-cysteine.On the other hand, two peaks are observed for two-layered sample.The position of the lower-energy peak (marked A) is close to that for the multi-layered sample.It is deduced that the higher-energy peak at 2480.4 eV (marked B) originates from the formation of Au-S configuration between sulfur and gold atoms.Two peaks are observed is important to know the surface structure.This result indicates that a part of two-layered L-cysteine surface formed some island structures.The energy shift of the S 1s between two-layered and multi-layered L-cysteine is 8.2 eV.We consider that sulfur atom's charge donation may be [δ + ] state.However it is difficult to explain the origin of this chemical shift between L-cysteine and Au, there is one possibility that the functional groups in amino acid influence to the energy shift.The detail of this discussion will be described later.Now we will discuss about the origin of the chemical shifts between two-layered and multi-layered films, observed both in NEXAFS and XPS.First, we describe the S 1s binding energies E B (S 1s) in XPS spectra.The S 1s chemical shift ∆E(S 1s) between two-and multi-layered films is given by where the subscripts "mono" and "multi" represent twohttp://www.sssj.org/ejssnt(J-Stage: http://www.jstage.jst.go.jp/browse/ejssnt/) layer and multi-layer, respectively.All of the energies are measured from the Fermi level of the gold substrate.
If we adopt a one-electron transition (or single particle) approximation, the resonance energy E(res) observed in NEXAFS spectrum is given by where E(un) is the energy of unoccupied states into which the core electrons are excited, and ∆R is the intra-atomic and extra-atomic relaxation energy due to the existence of the core-hole (core-hole effect).The value of E(un) mono is almost zero because of the alignment of the Fermi level at the interface between adsorbed cysteine and gold surface.The value of E(un) multi is nearly equal to E g /2, where E g is the energy-gap of solid L-cysteine.There is no reliable data about the energy-gap of multi-layered cysteine, but the value would be negligible in the present discussion because the film is fairly thin (less than 1.0 nm).Thus from Eqs. ( 3), ( 4) and ( 5), we obtain next equation.

E(res) mono −E(res
(6) As to the relaxation energy (∆R), it is reported that, in the case of weakly bound molecules, the excited electrons are pull down due to Coulomb interaction by the core hole [23].For chemisorbed molecules on metal surface, on the other hand, the excited electrons are screened by the metal substrate, so the relaxation energy becomes small compared to that of the weakly bound system, resulting in ∆R multi > ∆R mono .So we consider that the large energy separation in NEXAFS spectra in comparison with that in XPS is due to the large relaxation energy in multi-layer.
In order to confirm whether the unique chemical shift observed in two-layered L-cysteine on gold is specific only to S-Au bond or not, we have investigated the XPS and NEXAFS spectra for the other substrates.In this time, we select two kinds of substrate.One is indium tin oxide (ITO) substrate and the other is Cu one.ITO is on behalf of transparent oxide conductor and that substrate has been already used as organic device.Cu is one of the noble metals whose properties are similar to those of gold.Figures 3 (a), (b) and (c) show the S K-edge NEX-AFS spectra for two-layered L-cysteine on Au, ITO and Cu substrate, respectively.A sharp peak is observed at 2473.4.eV (marked A) at ITO and Cu substrate however there is no peak at 2480.4 eV attributed to the S-Au bond.According the Yagi's paper [24], the S 1s → σ * (S-C) resonance energy for L-cysteine on Cu is about 2472.6 eV, which is not so different from our data 2473.4.eV (peak A spectrum (c) of Fig. 3).The important result of the present paper is the finding of the specific peak in NEXAFS spectrum (peak B in spectrum (a) of Fig. 3) which is observed only S-Au interface between L-cysteine and gold surface.
Figure 4 shows the S 1s XPS spectrum of two-layered L-cysteine on Au and ITO substrates.The lower binding energy peak at 2472.4 eV belongs to L-cysteine adsorbed on ITO surface and there is no specific bond state between S and ITO substrate.
These results lead to important conclusion the observed specific chemical state representing large chemical shift is inherent to sulfur-gold interface.As to the S 2p XPS, Petoral Jr. et al. reported that the energy of the S 2p peak in XPS is lower by about 2 eV than as a result of a strong molecule-surface interaction in the S 2p XPS spectra for thiol-Au bond [25].Similar results were reported by the other groups [26][27][28][29].Castner et al. has also reported that the binding energy of S 2p of organic thiol and disulfide binding interactions with gold surfaces is the 163.2-163.5 eV and 162.0 eV, respectively [30].However we cannot compare our data with reference data directly, because x-ray source, molecular structure and technical method to prepare the mono-layered films are not the same condition.In the present work, it should be noted that the direction of the energy shift observed in S 1s XPS is opposite to those reported for the other http://www.sssj.org/ejssnt(J-Stage: http://www.jstage.jst.go.jp/browse/ejssnt/) e-Journal of Surface Science and Nanotechnology S-metal systems observed in S 2p XPS.Considering the opposite direction of the present chemical shift, it is presumed that the charge transfer provides from sulfur to gold for two-layered L-cysteine.Another evidence for the positive charge state of sulfur is obtained from the difference between XPS and NEXAFS spectra for two-layered sample.For two-layered sample, the S 1s peak in XPS has double peak structure (spectrum (b) of Fig. 2), but only one peak is seen in the NEXAFS (spectrum (b) of Fig. 1).This difference can be interpreted by the orbital population in the S 3p* states as follows.
In the present experiment, the XPS and NEXAFS measurements were carried out on the same surface.The Kshell spectra of atoms and molecules contain a variety of pronounces resonance which corresponds to electronic transitions from a K-shell to valence unoccupied states near the vacuum level and therefore fall at an excitation energy close to the ionization potential (IP).In the case of S K-edge excitation, the intensity of the S 1s → σ*(S-C) resonance is enhanced when S 3p* orbital becomes empty.For XPS, intensity of the photoelectrons is determined only by the photoelectron cross section of core levels.In the present case, our data show that the sulfur atoms at the S-Au interface represent likely [δ + ] state on the basis of the XPS peak energy.Thus, the valence electrons (mainly composed of S 3p orbitals) in sulfur atoms are donated to the gold substrate.This means that the orbital population in the S 3p states seems to be an empty.Therefore, the intensity of the 2484.0eV peak in the NEX-AFS spectra corresponding to the dipole transition from the S 1s to unoccupied S 3p* state is enhanced.We consider this is the reason why the XPS from the two-layer film includes two features characteristic but the same effect is not seen in the NEXAFS.
We assumed that functional groups of amino acids participate in strong bond formation about Au-S interface.In these circumstances, we conclude that the observed unique chemical shift interpreted by the electron donation from sulfur to gold is inherent to sulfur-containing amino acids.This specific bond gives important information to know the influence from adsorption bio-molecules in construction of the new bio-molecular devices.

IV. CONCLUSIONS
We have measured the S K-edge NEXAFS and XPS spectra for adsorbed L-cysteine on gold as well as copper and ITO surfaces.For multi-layered L-cysteine film, the S 1s → σ*(S-C) resonance peak was observed in the NEXAFS spectra at 2475.0 eV.While for two-layered Lcysteine film, this resonance peak was observed at 2484.0 eV, which is higher by 9.0 eV than that for multi-layered L-cysteine.In addition, in the S 1s XPS spectra for twolayered L-cysteine film, newly one peak was observed at 2480.4 eV, which is higher binding energy side by 8.2 eV than that for multi-layered one.On the other hand, high-energy peaks observed in NEXAFS and XPS for Lcysteine-gold system were not observed for L-cysteine on Cu and ITO substrates.These result indicated that Au-S bond is specific bonding compare with Cu and ITO substrates.Furthermore, the direction of the S 1s energy shift for Au-S bond configuration is opposite to that for the normal metal sulfides.This means that the electrons are donated from the sulfur atom to the gold substrate, and the sulfur atoms are positively charged at likely [δ + ] states.We conclude that the strong Au-S bonds that are recently used for the immobilization of molecules are due to the observed specific Au-S bond between L-cysteine and Au surface.

FIG. 1 :
FIG. 1: S K-edge NEXAFS spectra for L-cysteine taken by total electron yield.The spectra (a) and (b) represent multilayered and two-layered samples, respectively.

FIG. 2 :
FIG.2: S 1s XPS spectra for L-cysteine on the Au surface which was taken at hν = 3000 eV.The spectra (a) and (b) represent multi-layered and two-layered samples, respectively.The solid Gaussian curves show the components that were used to fit the experimental data.

FIG. 3 :
FIG. 3: S K-edge NEXAFS spectra for two-layered L-cysteine taken by total electron yield.The spectra (a), (b) and (c) represent Au substrate, ITO substrate and Cu substrate sample, respectively.