Adsorption Reaction of Amino Acid Molecule on Pd Thin Layer Surface Constructed by Nano-dots under Water Environment

The adsorption reaction of L-cysteine on Pd thin layer surface constructed by nano-dots under water environment has been investigated by S K-edge NEXAFS with He-path system under atmospheric pressure. It is revealed that L-cysteine physisorbs on Pd/Al2O3/NiAl(111) substrate surface through the thiol group and the carboxyl one, and chemisorbed L-cysteine exists on the surface as L-cysteine thiolate. Before and after dipping the Pd/Al2O3/NiAl(111) substrate into L-cysteine aqueous solution, the surface morphology is observed using AFM. The morphology of the Pd/Al2O3/NiAl(111) substrate is composed of many nano-dots before dipping. After dipping, the nano-dots are enlarged. [DOI: 10.1380/ejssnt.2011.289]


I. INTRODUCTION
Active oxygen is one of the origins of both aging and carcinogenesis in our body.Platinum group metals have an advantage removing the active oxygen.The cosmetic and pharmaceutical products containing nanoparticles of platinum group metals have been already marketed in our surroundings.However, the safety of ingested nanoparticles in our body has not been demonstrated enough.Therefore, it is of great importance to reveal the mechanism of adsorption reaction between those nanoparticle surfaces and biomolecules in vivo.
Approximately 70% of our body consists of water.To clarify the reaction between the metal nanoparticles and the biomolecules in vivo, there is a need to measure the reaction under water environment.Many amino acid molecules exist in our body.L-cysteine, which is one of the amino acids, has been selected in our study because it has a thiol group with a high reactivity on the transition metal surface.Yagi et al. have cleared that the L-cysteine has chemisorbed on polycrystalline nickel or cupper surface, and a part of adsorbed molecules has been dissociated by the method of S K-edge Near-edge X-ray Absorption Fine Structure (NEXAFS) measurements [1].Although they have focused on solid-liquid interface between transition metal surface and L-cysteine aqueous solution, the S K-edge NEXAFS measurements have been carried out under vacuum condition.To make in vivo condition, we have decided to observe the adsorption reaction of those samples under water environment.
Recently, we have investigated the molecular adsorption system on nanoparticle surfaces of platinum group metals under water environment by means of S K-edge NEXAFS with He-path system under atmospheric pressure.For example Gohda et al. have fabricated Rh(PVP) nanoparticles that are synthesized by reducing Rh ion in ethanol-water solvent with water-solution polymers of PVP (polyvinylpyrrolidone), and added L-cysteine powder to the nano-colloidal solution [2,3].They have revealed an adsorption reaction between Rh nanoparticles and the L-cysteine under water environment.As a result, adsorbed L-cysteine on Rh nanoparticle is dissociated into L-cysteine thiolate.However, it has possibilities to observe a reaction between PVP and the L-cysteine because PVP exists around Rh nanoparticles as an impurity.Thus, it needs to fabricate nanoparticles with clean surfaces so that we want to reveal only the adsorption reaction between nanoparticles and the L-cysteine.To fabricate nanoparticles with clean surface, we have used the gas evaporation method with noble gas [4,5].Fabrication of Pd nanoparticles, which belongs to the platinum group metals, was successful with the method with He in our previous studies [6,7].Therefore, we have focused on the adsorption reaction between Pd nanoparticles and the L-cysteine under water environment.
As a first step, we study the adsorption reaction of Lcysteine on Pd thin layer surface constructed by Pd nanodots under water environment by S K-edge NEXAFS.
The NiAl(111) single crystal was polished with 0.05 µm Al 2 O 3 paste to obtain a mirror finish.The impurities on the NiAl(111) single crystal surface were removed by Ar + ion sputtering (3 keV, 2 µA, for 1 hour) in an ultrahigh vacuum chamber.
To create well-ordered ultrathin α-Al 2 O 3 on the NiAl(111) substrate, we used some parameters reported by Nomoto et al. [8].The NiAl(111) substrate was heated at 600 K in oxygen environment and annealed at 1000 K for 20 min.A Pd thin layer of 0.1 ML was deposited on the α-Al 2 O 3 /NiAl(111) substrate by an electron beam evaporator equipped with Pd wire (purity: 99%).
L-cysteine aqueous solution (0.25 mol/l) was prepared by mixing the distilled water and L-cysteine powder purchased from SIGMA-ALDRICH.Prepared substrates were dipped into L-cysteine aqueous solution for 12 hours at room temperature.Those substrates were taken out of the solution and rinsed with distilled water.To keep the condition of water environment and to obtain the NEX-AFS spectra under water environment in S K-edge energy region, those substrates were wrapped with polyethylene film of 12 µm thickness.After finishing the NEXAFS measurement under water environment, all substrates were rinsed again and dried in air.The NEXAFS measurements for the dried substrates were also carried out.As a standard spectrum, L-cysteine aqueous solution injected with a polyethylene cell was measured.To obtain a background spectrum for S impurities on the NiAl(111) substrate surface, moreover, the substrate without dipping into L-cysteine aqueous solution was measured.
To reveal morphology of the Pd/α-Al 2 O 3 /NiAl(111) substrate surface for both before and after dipping into L-cysteine aqueous solution, AFM observation was carried out by means of NanoScope III-a (Veeco Instruments) with tapping mode.S K-edge NEXAFS measurements were done by fluorescence X-ray yield method using the atmospheric XAFS measurement system with He-path at the BL-3 on Hiroshima Synchrotron Radiation Center (HSRC) [9,10].The incident X-ray energy was calibrated on the assumption that the first peak of K 2 SO 4 appears at 2481.70 eV.The fluorescence yield detection was employed using a gas-flow type proportional counter with P-10 gas (10% CH 4 in Ar).The energy region of NEXAFS measurement was 2467-2489 eV.Energy step and dwell time were 0.2 eV and approximately 120 seconds/point, respectively.

III. RESULTS AND DISCUSSIONS A. AFM observation
To reveal morphology for the fabricated the Pd/Al 2 O 3 /NiAl(111) substrate surface, the surface is observed with AFM.The AFM image is shown in Fig. 1.The many nano-dots are observed in the circles shown in Fig. 1.The average diameter of those nano-dots is estimated to be 1.2±0.2nm.In previous study, it is also reported that the morphology for the Pd evaporated layer surface is composed of nano-dots [11].The surface of the Pd/Al 2 O 3 /NiAl(111) substrate, which is dipped into L-cysteine aqueous solution for 12 hours, is subsequently observed with AFM.The result is shown in Fig. 2. Enlarged nano-dots are can be seen.The average diameter of those enlarged nano-dots is estimated to be 2.5±0.4 nm.The following three reasons can be thought of to be responsible for the enlargement of average diameter for nano-dots: i) Reaction between Pd nano-dots and L-cysteine under water environment, ii) Air oxidation of Pd nano-dots, and iii) Diffusion and aggregation of Pd nano-dots on the Al 2 O 3 /NiAl(111) substrate under water environment.
It is necessary to reveal the dominant reason.To explain the enlargement of the nano-dots for each substrate surface, we will investigate the reaction for the Pd/Al 2 O 3 /NiAl(111) substrate surface under various conditions in the future.strates under water environment.Those substrates are dipped into L-cysteine aqueous solution for 12 hours.To identify a chemisorbed adsorbate on each substrate surfaces, the assignation of NEXAFS peak positions is necessary.From the results of previous studies, we have judged the peak positions of L-cysteine thiolate and atomic S [2,8,12,13].The spectra for L-cysteine aqueous solution and the NiAl(111) substrate without dipping into L-cysteine aqueous solution under dry environment are also shown as standard and background.The peak positions for L-cysteine aqueous solution and the NiAl(111) substrate are 2472.7 eV and 2481.9 eV, respectively.If the peaks in the energy region about 2470.5-2472.6 eV and at 2470.0 eV appear, it is possible that L-cysteine thiolate and atomic S created by the dissociation from L-cysteine thiolate exist on the substrate surface.When we pay attention to NEXAFS spectrum for the Pd/Al 2 O 3 /NiAl(111) substrate, the peak is located around 2472.7 eV and the peak width grows in comparison with that of the spectrum for L-cysteine aqueous solution.Therefore, it is found that L-cysteine is physisorbed on the Pd/Al 2 O 3 /NiAl(111) substrate surface and the chemisorbed L-cysteine exists as an adsorbate of L-cysteine thiolate.On the other hand, a small peak is shown at 2472.surface also.When we focus on the peak intensity of the L-cysteine on both substrates, the peak intensity for the Pd/Al 2 O 3 /NiAl(111) is much bigger than that for the Al 2 O 3 /NiAl(111).In addition, from the point of view for the automobile exhaust gas catalyst, Dohmae has reported that Pd-S bonding is stable in comparison with Rh-S one because Pd-S bonding energy is larger than that for Rh-S one [14].Hence, it is thought that most of L-cysteine molecules adsorb on the active Pd surface selectively, not on the inactive Al 2 O 3 surface.Moreover, a peak exists in the energy region of 2475-2485 eV for both substrates.Nomoto et al. have reported that this peak is assigned to SO x (x = 3, 4) [8].The peak positions for the Pd/Al 2 O 3 /NiAl(111) and the Al 2 O 3 /NiAl(111) substrates in the energy region of 2475-2485 eV are 2480.8eV and 2481.9 eV, which is the same as the peak position of the NiAl(111) substrate.In this study, it is thought that the SO x on the Pd/Al 2 O 3 /NiAl(111) substrate is formed by the coordinate bond between S of L-cysteine thiolate or atomic S and oxygen of H 2 O under water environment, and SO x on the Al 2 O 3 /NiAl(111) substrate is derived from the impurity of the NiAl(111) substrate.without dipping into L-cysteine aqueous solution is also shown as a background.Because the spectrum for the Pd/Al 2 O 3 /NiAl(111) substrate has the peak with broad width at 2472.7 eV, it is revealed that both L-cysteine and L-cysteine thiolate exist on the Pd/Al 2 O 3 /NiAl(111) substrate surface under dry environment, which is same as the result of the adsorption reaction under water environment.However, Yagi et al. have reported that Lcysteine dissociates on some metals surfaces under dry environment [1].Judging from those results, it can be thought that L-cysteine is not physisorbed on the Pd/Al 2 O 3 /NiAl(111) substrate surface as written in Section III.B.1, but chemisorbed on the surface through a thiol group and a carboxyl one.Because the peak assigned to L-cysteine is not observed in the spectrum for the Al 2 O 3 /NiAl(111) substrate under dry environment, on the other hand, it can be speculated that L-cysteine is physisorbed on the Al 2 O 3 /NiAl(111) substrate surface under water environment.

Under dry environment
Focusing on the energy region of 2475-2485 eV, the peaks assigned to SO x can be recognized in the spectra for both the Pd/Al 2 O 3 /NiAl(111) and the Al 2 O 3 /NiAl(111) substrates under dry environment.The peak position located at 2481.9 eV for the Pd/Al 2 O 3 /NiAl(111) substrate is similar to that for the NiAl(111) substrate.Therefore, it is thought that the peak for the Pd/Al 2 O 3 /NiAl(111) substrate is originating from the NiAl(111) substrate, which is including SO x on the surface as an impurity.The peak intensity at 2480.8 eV for the Pd/Al 2 O 3 /NiAl(111) substrate under dry environment is reduced in comparison with that for the same substrate under water environment.Thus, it can be thought that the coordinated H 2 O desorbs from both S of L-cysteine thiolate or atomic S on the Pd/Al 2 O 3 /NiAl(111) substrate surface.If we compare the peak intensity at 2480.8 eV for the Al 2 O 3 /NiAl(111) substrate under dry environment with the peak intensity under water environment, the intensity under dry environment is bigger than the intensity under water environment.Under dry environment, moreover, there is no peak at 2472.7 eV assigned to physisorbed Lcysteine for the Al 2 O 3 /NiAl(111) substrate.Because of those results, it is speculated that physisorbed L-cysteine on the Al 2 O 3 /NiAl(111) substrate surface is changed into SO x by bonding with oxygen in air.We focus on the peak intensity located at 2481.9 eV for the three spectra shown in Fig. 4. Owing to the deposition layer of Pd and/or Al 2 O 3 on the NiAl(111) substrate, those intensities become small in order of the Pd/Al 2 O 3 /NiAl(111), the Al 2 O 3 /NiAl(111) and the NiAl(111) substrates under dry environment.

IV. CONCLUSIONS
The morphology of the Pd/Al 2 O 3 /NiAl(111) substrate surface is observed with AFM.As the result, many nano-dots are observed on the surface.
When the Pd/Al 2 O 3 /NiAl(111) substrate is dipped into L-cysteine aqueous solution, it is clear that the nano-dots are enlarged.The adsorption reaction of L-cysteine on Pd thin layer under water environment has been investigated by S K-edge NEXAFS measurement.L-cysteine physisorbs on the Pd/Al 2 O 3 /NiAl(111) substrate through the thiol group and the carboxyl one under water environment, and chemisorbed L-cysteine on the substrate exists as the adsorbate of L-cysteine thiolate.It is found that L-cysteine molecules adsorb on the Pd surface selectively under water environment.

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FIG. 1: AFM image of the surface morphology for the Pd/Al2O3/NiAl(111) substrate before dipping into L-cysteine aqueous solution.Many nano-dots are observed in the circles.

Figure 4
Figure 4 shows S K-edge NEXAFS spectra for the Pd/Al 2 O 3 /NiAl(111) and the Al 2 O 3 /NiAl(111) substrates measured under dry environment.Those substrates, which are same as the substrates shown in Fig. 3, are dried in air.The spectrum for the NiAl(111) substrate