Self‐assembled monolayers of mercaptosuccinic acid monolayers on silver and gold surfaces designed for protein binding. Part II: vibrational spectroscopy studies on cytochrome c immobilization

An attempt has been made for using MSA‐modified electrodes as linkage monolayers for electrostatic and covalent binding of cytochrome c (Cc). For MSA monolayers grown from an aqueous solution on Ag, attachment of Cc in its native state is proved in the case of covalent bonding. Electrostatic immobilization of Cc at pH 7 results in presence of at least some amount of Fe²⁺ high‐spin configuration and/or Fe³⁺ oxidation state. Native protein Fe²⁺ low‐spin state of Cc is observed after applying a negative potential to the Ag electrode. The influence of the solvent used for the preparation of the MSA monolayer and thiol surface coverage of the Ag surface was studied. It was shown that the key factor to obtain the native structure of Cc is the successful blocking of the metal surface by the MSA linking layer. IRRAS measurements of MSA on monocrystalline gold (111) at neutral pH confirm the successful electrostatic Cc immobilization, which preserves the Fe²⁺ oxidation state of the chromophore on this substrate. Copyright © 2007 John Wiley & Sons, Ltd.     

Out‐of‐plane deformations of the heme group in different ferrocytochrome c proteins probed by resonance Raman spectroscopy

We measured the low‐wavenumber polarized resonance Raman spectra of horse heart (hhc), chicken (chc) and yeastC102T (yc) ferrocytochromes c with Soret excitation. We examined the out‐of‐plane (oop) deformations of the heme groups by virtue of relative intensities and depolarization ratios of a variety of oop and in‐plane (ip) Raman active bands. Analysis of relative Raman intensities shows differences in deviation from planarity of the heme groups of yeast, horse heart and chicken cytochromes c. The heme groups in cytochrome c proteins have been shown by normal coordinate static deformation (NSD) analysis from crystal structures to exhibit a dominant ruffling (B_1u) deformation. As a consequence the B_1u modes, γ₁₀ − γ₁₂, become resonance Raman active. We used normalized Raman intensity ratios and depolarization ratios of oop Raman active modes, whose intensities are attributable to specific nonplanar deformations, to estimate and compare their Franck‐Condon‐type and Jahn‐Teller‐type coupling magnitudes for horse heart, chicken and yeast ferrocytochrome c at neutral pH. These coupling magnitudes allow for a quantitative comparison of oop deformations between individual heme groups. Chicken ferrocytochrome was found to have the largest ruffling deformation of the three investigated proteins, followed by horse heart and yeast cytochrome c. The heme group of the former is slightly more ruffled than the corresponding active site of the latter, while saddling in both proteins is substantially larger than in chicken ferrocytochrome c. The Raman data are sensitive enough to allow a comparison of lesser deformations. Doming, which is a kinetic coordinate in many heme proteins, is largest in chicken and smallest in yeast cytochrome c. Waving is largest in yeast, followed by horse heart and chicken cytochrome c. Propellering deformations could be compared for chicken and horse heart cytochrome c and were found to be substantially larger in the latter. A comparison with heme deformations obtained from X‐ray structures (for horse heart and yeast cytochrome c) and from molecular dynamics simulations (MDS) (performed for all three proteins) yields some agreement with the main ruffling and saddling deformations derived from the crystal structures, whereas the heme conformations produced by MDS seem to account better for smaller deformations like doming and propellering. The present study demonstrates the usefulness of resonance Raman spectroscopy for the analysis of nonplanar deformations in heme proteins. Copyright © 2008 John Wiley & Sons, Ltd.     

Comparison of SERRS and RRS excitation profiles of [Fe(tpy)2]2+ (tpy = 2,2':6',2'‐terpyridine) supported by DFT calculations: effect of the electrostatic bonding to chloride‐modified Ag nanoparticles on its vibrational and electronic structure

Nonresonance (or normal) Raman scattering (NRS), resonance Raman scattering (RRS), surface‐enhanced Raman scattering (SERS), and surface‐enhanced RRS (SERRS) spectra of [Fe(tpy)₂]²⁺ complex dication (tpy = 2,2':6',2''‐terpyridine) are reported. The comparison of RRS/NRS and SERRS/SERS excitation profiles of [Fe(tpy)₂]²⁺ spectral bands in the range of 445–780 nm is supported by density functional theory (DFT) calculations, Raman depolarization measurements, comparison of the solid [Fe(tpy)₂](SO₄)₂ and solution RRS spectra, and characterization of the Ag nanoparticle (NP) hydrosol/[Fe(tpy)₂]²⁺ SERS/SERRS active system by surface plasmon extinction spectrum and transmission electron microscopy image of the fractal aggregates (D = 1.82). By DFT calculations, both the Raman active modes and the electronic states of the complex have been assigned to the symmetry species of the D_2d point group. It has been demonstrated that upon the electrostatic bonding of the complex dication to the chloride‐modified Ag NPs, the geometric and ground state electronic structure of the complex and the identity of the three different metal‐to‐ligand charge transfer (¹MLCT) electronic transitions remain preserved. On the other hand, the effect of ion pairing manifests itself by a slight change in localization of one of the electronic transitions (with max. at 552 nm) as well as by promotion of the Herzberg–Teller activation of E modes resulting from coupling of E and B₂ excited electronic states. Finally, the very low, 1 × 10⁻¹¹ M SERRS spectral detection limit of [Fe(tpy)₂]²⁺ at 532‐nm excitation is attributed to a concerted action of the electromagnetic and molecular resonance mechanism, in conjunction to the electrostatic bonding of the complex dication to the chloride‐modified Ag NP surface. Copyright © 2014 John Wiley & Sons, Ltd.     

Electrochemical and in situ SERS spectroelectrochemical investigations of 4‐methyl‐4H‐1, 2, 4‐triazole‐3‐thiol monolayers at a silver electrode

Electrochemically anticorrosive behavior of 4‐methyl‐4H‐1, 2, 4‐triazole‐3‐thiol (MTTL) self‐assembled monolayers (SAMs) on the silver electrode was studied by means of electrochemical impedance spectroscopy (EIS) and polarization measurements. The promising inhibition effect of the MTTL for silver had been affirmed. Results of surface‐enhanced Raman scattering (SERS) experiments indicated that the MTTL molecule in a tilted orientation was self‐assembled on the silver surface through S⁶ and N² atoms to form monolayers. An in situ electrochemical SERS experiment implied the changes of adsorption fashion of MTTL momolayers on the silver surface with the potential shifted to more negative direction. Copyright © 2009 John Wiley & Sons, Ltd.     

The influence of pH and anions on the adsorption mechanism of rifampicin on silver colloids

The influence of pH and anions on the adsorption mechanism of rifampicin on colloidal silver nanoparticles has been analysed by electronic absorption, resonance Raman (RR) and surface‐enhanced resonance Raman spectroscopy (SERRS). Rifampicin is a widely used antibiotic with a zwitterionic nature. SERRS spectra of rifampicin adsorbed on silver sols, prepared using hydroxylamine hydrochloride as reducing agent, undergo dramatic changes upon lowering the pH. The spectral form changes progressively from that characteristic of chemisorbed rifampicin (at pH > 7) to one very similar to the rifampicin RR spectrum (at lower pH), indicative of a modification of the adsorption mechanism on the surface of the Ag nanoparticles. The RR‐type SERRS spectrum is proposed to result from formation of an ion pair between rifampicin and Cl⁻ anions, which, deriving from the colloid preparation, are adsorbed on the Ag surface. The addition of anions to the hydroxylamine hydrochloride sol facilitates conversion from the chemisorbed to ion pair form and leads to an order of magnitude increase in the SERRS signal. Copyright © 2007 John Wiley & Sons, Ltd.     

Spacer‐free SERRS spectra of unperturbed porphyrin detected at 100 fM concentration in Ag hydrosols prepared by modified Tollens method

Surface‐enhanced resonance Raman scattering (SERRS) spectroscopy is a powerful tool for the selective and sensitive detection of porphyrins. We report a repeatable detection of unperturbed free base 5,10,15,20‐tetrakis(1‐methyl‐4‐pyridyl)21H,23H‐porphine in concentrations as low as 1 × 10^–13 M in the final system when Ag nanoparticles prepared by modified Tollens method are employed as Raman scattering amplifiers, i.e. without any further post‐modifications of their surface. Most probably the residues of sugar oxidation present on Ag nanoparticle surface are responsible for the effective protection of 5,10,15,20‐tetrakis(1‐methyl‐4‐pyridyl)21H,23H‐porphine molecules against the metalation by silver. Copyright © 2011 John Wiley & Sons, Ltd.     

Growth kinetics of self-assembled monolayers of thiophene and terthiophene on Au(111): An infrared spectroscopic study

The growth kinetics of self-assembled monolayers (SAMs) of thiophene compounds on Au(111) surfaces was revealed by Fourier-transform infrared reflection absorption spectroscopy (FT-IR-RAS). Thiophene and terthiophene form well-ordered SAMs on Au(111) surfaces by immersing gold substrates into their ethanol solutions for ca. 15 h. Gibbs free energies for the adsorption processes of thiophene and terthiophene were found to be identical. However, the growth and molecular orientation of SAMs are different between two thiophene compounds. Terthiophene in SAMs orients parallel to the surface. The SAM growth of terthiophene obeys a time-dependent Langmuir scheme. On the other hand, the thiophene SAM undergoes a two-step growth process with unique molecular orientations. In the primary phase, thiophene assumes a parallel orientation on the Au(111) surface. In the second phase, thiophene is oriented close to the normal of the surface. The different growth process between thiophene and terthiophene is attributable to the topology of sulfur positions in the molecules. Received 23 May 2001 and Received in final form 11 February 2002     

1H NMR Studies of 1-Ethylimidazole Binding to Cytochrome C

Abstract

Kinetic and equilibrium data for 1-EtIm binding to cyt c at temperature range of 303–319K have been determined at pH 7.0 by using ¹HNMR method. Thermodynamic values (ΔH°=39.5 kJ mol^?1, ΔS° = 154 J mol^?1K^?1 and Ea = 142 kJ mol^?1) are obtained from Van't Hoff and Arrhenius's relations. Some hyperfine shifted resonances of l-EtIm cyt c have been assigned for the first time using 1D saturation transfer experiments. The origin of the asymmetric spin density distribution in heme groups of 1-EtIm cyt c and the reason of low affinity of cytochrome c for 1-EtIm are also discussed move toward upfield from original position. However, Te-2, 4 substituents in cyt c are more electron withdrawing than the propionic acid side chains, which lead to the 5-methyl group going toward downfield.     

A stable ‘sandwich’ system of Surface‐Enhanced Resonance Raman Scattering for the analysis of β‐carotenes in a photosynthetic pigment‐protein complex

In plants, Photosystem I (PSI) is composed of a core complex and a membrane‐associated antenna complex light‐harvesting complex I that captures light and funnels its energy to the core complex. To obtain Raman structural information on β‐carotenes embedded in the PSI core complex, a ‘sandwich’ system of roughened silver slice: target protein complexes: single silver nanoparticles was fabricated for Surface‐Enhanced Resonance Raman Scattering (SERRS) measurements. This study provided a method to overcome spectral irreproducibility, which is the main drawback of Surface‐Enhanced Raman Scattering/SERRS‐based studies. The Raman spectra of β‐carotenes embedded in the PSI core complex can be obtained at very low sample concentrations (1–5 µg Chl/ml) and high signal/noise ratios. The β‐carotenes in the spinach PSI core complex were predominantly all‐trans configuration. The membrane protein‐mediated adsorption of silver nanoparticles induced the uniform distribution of a large number of single nanoparticles, which contributed to achieving highly reproducible SERRS spectra. This study is the first to apply single silver nanoparticle‐based SERRS analysis in membrane proteins. Copyright © 2013 John Wiley & Sons, Ltd.     

pH‐dependent surface‐enhanced Raman scattering studies of N‐acetylalanine monolayers self‐assembled on a silver surface

Monolayers of N‐acetylalanine on a metallic surface can serve as a biocompatible functional interface to construct biosensors. In the present paper, the surface‐enhanced Raman scattering (SERS) spectra of N‐acetylalanine monolayers self‐assembled on a silver surface under different pH were recorded. Assignments of the obtained spectra were carried out by density functional theory (DFT) calculations (BLYP/6‐311G). On the basis of the SERS effect, the nature of adsorption of N‐acetylalanine on a silver surface was deduced. It can be concluded that the fully protonated N‐acetylalanine is adsorbed on the silver surface via the imine group together with the carboxylate group, while it anchored onto the surface not only through both the imine and the carboxylate groups but also through the amide group after being completely deprotonated in the basic solution. Copyright © 2007 John Wiley & Sons, Ltd.     

Interfacial kinetic enhancement of metal ion adsorption on binary mixed self-assembled monolayers

The adsorption of metal ions, a type of surface reaction on binary mixed self-assembled monolayers (SAMs) on a gold surface composed of 1,6-hexanedithiol (HDT) with 11-mercaptoundecanoic acid (MUA), was monitored by in situ surface plasmon resonance (SPR) measurements. The differential SPR reflectance (ΔR) enables the kinetics of adsorption of Pt²⁺ on the mixed SAMs to be investigated. Unlike single HDT SAM, kinetic analyses of the mixed SAMs showed that the rate of adsorption of Pt²⁺ was enhanced and that it was highly dependent on the fraction of MUA present. These SPR measurements suggest that the adsorption rate of metal ions can be readily manipulated simply by using mixed SAMs.     

Silica‐overcoated substrates for detection of proteins by surface‐enhanced Raman spectroscopy

Ag film over nanosphere (AgFON) substrates for surface‐enhanced Raman spectroscopy (SERS) are shown to be ineffective for the detection of proteins in phosphate buffer solution (PBS) because of the decomposition of the substrate resulting in a total loss of SERS activity. However, modification of these substrates with SiO₂ overlayers overcomes this problem. The SiO₂ overlayers are produced by filtered arc deposition (FAD) and are characterised by atomic force microscopy (AFM). Their porosity is examined using Raman spectroscopy and the detection of cytochrome c and bovine serum albumin in PBS is successfully demonstrated. These findings show promise for the detection of proteins in biologically relevant conditions using Ag‐based SERS substrates. Copyright © 2008 John Wiley & Sons, Ltd.     

Trans‐2‐Aminocyclohexanol derivatives as pH‐triggered conformational switches

A series of trans‐2‐aminocyclohexanol derivatives have been explored as powerful conformational pH triggers. On protonation of the amino group, a conformer with equatorial position of ammonio and hydroxy groups becomes predominant because of an intramolecular hydrogen bond and electrostatic interactions. The energy of these interactions was estimated to be above 10 kJ/mol and in some models exceeded 20 kJ/mol (strong enough to twist a ring in tert‐butyl derivatives). As a result of this conformational flip, all other substituents are forced to change their orientation. If the substituents are designed to perform certain geometry‐dependent functions, for example, as cation chelators or as lipid tails, such acid‐induced transition may be used to control the corresponding molecular properties. The pH sensitivity of conformational equilibria was explored by ¹H nuclear magnetic resonance spectroscopy (NMR), and the titration curves were used for estimation of the pK_a values of protonated compounds that varied from 2.6 to 8.5 (in d₄‐methanol) depending on the structure of amino group. Thus, trans‐2‐aminocyclohexanols can be also used as conformational pH indicators in organic solvents.     

Label‐free detection of binary mixtures of proteins using surface‐enhanced Raman scattering

The surface‐enhanced Raman scattering (SERS) spectra of five binary mixtures of proteins were studied, including mixtures of apomyoglobin (apMb) and lysozyme, insulin and lysozyme, Mb and cytochrome c (cyt c), Mb and apoMb, and cyt c and human serum albumin. For mixtures in which both proteins did not contain chromophores, the variations in the intensities of SERS signals with changes in the relative concentration of the two proteins were largely proportional to the relative concentration of the protein mixtures. Similar results were obtained for mixtures in which both the proteins contained chromophores. However, for mixtures in which one protein contained chromophores and the other did not, the SERS intensities of the protein mixtures showed a bell‐shaped variation with changes in the compositions of the mixtures. The present study provides new insights into the application of SERS spectroscopy to label‐free detection in a binary of mixture of proteins. The complexities in both SERS phenomena and protein structures should be considered in the analysis of the SERS spectra of protein mixtures. By monitoring the heme band of a protein containing a heme and the NO₃^– band of a protein without a heme, the simultaneous detection of both proteins in a binary mixture may be possible. It has also been found that the larger the size of the protein, the smaller is the SERS intensity of that protein. Copyright © 2011 John Wiley & Sons, Ltd.     

An atomic force microscope study of vanadium-benzene sandwich clusters soft-landed on self-assembled monolayers

Multiple-decker vanadium-benzene sandwich clusters V_n(benzene)_n+1 produced by a laser-vaporization synthesis method were soft-landed onto self-assembled monolayers of alkanethiol (C₁₈H-SAM) and fluorinated alkanethiol (C₁₀F-SAM) at 200 K. Noncontact atomic force microscopy has been used to examine the resulting adsorption states of the clusters landed on the SAMs at room temperature. For each SAM substrate, the aggregates of the deposited clusters were observed at the vacancy islands and near the steps of the SAM surface. The result indicates that, at room temperature, the clusters landed on the SAM substrate thermally diffuse on the surface to form columnar-shape islands around the defect sites of the SAM surface.     

Surface‐enhanced Raman scattering study of monolayers formed from mixtures of 4–mercaptobenzoic acid and various aromatic mercapto‐derivative bases

Monolayers formed from aromatic compounds have many potential applications, for example in construction of bioelectronic elements having high efficiency of electron transfer. In this paper, the composition of monolayers formed on silver surfaces from mixtures of 4‐mercaptobenzoic acid (MBA) and four model (stable and easily available) aromatic thiols with strong base properties: 4,6‐diamino‐2‐mercaptopyrimidine (APY), 1H‐1,2,4‐triazole‐3‐thiol (HTR), 4‐methyl‐4H‐1,2,4‐triazole‐3‐thiol (MTR) and 3‐amino‐1,2,4‐triazole‐5‐thiol (ATR), were determined from surface‐enhanced Raman scattering (SERS) measurements. Our investigations showed that among studied aromatic bases, APY is the most promising candidate for the formation of mixed monolayers with MBA. In the whole pH range studied (2–12.5), mixed MBA + APY monolayers with similar surface concentration of both components are formed during the adsorption from the 0.5 mM MBA + 0.5 mM APY aqueous solution. Desorption of MBA and APY from the mixed monolayer is, however, significantly different. During immersion in water, surface concentration of APY decreases significantly faster than MBA (a significant part of the adsorbed MBA molecules is present on the silver surface even after 2 h of soaking in water). Presence of chlorides, bovine serum albumin or laccase in the surrounding solution does not observably influence the structure of MBA + APY monolayers. The properties of monolayers formed from MBA and substituted triazoles were found to be significantly different than those of MBA + APY monolayers. Copyright © 2009 John Wiley & Sons, Ltd.     

Potential‐induced Raman behavior of individual (R)‐di‐2‐naphthylprolinol molecules on a Ag‐modified Ag electrode

The applicability of surface‐enhanced Raman spectroscopy is demonstrated to probe the adsorption behavior of individual molecules on a Ag electrode. High‐quality SERS spectra of (R)‐di‐2‐naphthylprolinol (DNP) were obtained from ultradilute solutions (10⁻¹² M ) on the Ag‐nanoparticle‐modified Ag electrode, which is attributed to the high electromagnetic (EM) effect of the SERS‐active system as well as to the strong adsorption and interaction of DNP molecules with Ag. The stable SERS spectra present remarkable potential dependence, which gives evidence for the behavior of individual DNP molecules on the Ag surface. Based on statistical analysis for the probability of DNP molecules located in ‘hot spots’, we propose an SERS mechanism for individual molecules in the electrode system, in combination with the hot‐spot model and orientation of the probe molecules. Copyright © 2010 John Wiley & Sons, Ltd.     

Fabrication of 6,13‐bis(triisopropyl‐silylethynyl)–pentacene thin‐film transistors with the silver ink transfer method using a polymer stamp

We fabricated 6,13‐bis(triisopropylsilylethynyl)–pentacene (TIPS–pentacene) thin film transistors using a direct metal transfer method. Using different metals, such as Au and Ag ink, electrode patterns are formed from the relief region of the polymer mold. TIPS–pentacene TFTs using the Ag ink transfer method show a similar performance to those using the Au metal transfer method. This method has advantages over the Au metal transfer method because it does not require vacuum equipment and a dry etching process. The self‐assembled monolayer (SAM) treated device exhibits a carrier mobility of 9.5 × 10^–2 cm²/V · s, and an on/off ratio of 4.6 × 10⁴. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Surface modification of cobalt-chromium-tungsten-nickel alloy using octadecyltrichlorosilanes

Cobalt-chromium (Co-Cr) alloys have been extensively used for medical implants because of their excellent mechanical properties, corrosion resistance, and biocompatibility. This first time study reports the formation and stability of self-assembled monolayers (SAMs) on a Co-Cr-W-Ni alloy. SAMs of octadecyltrichlorosilanes (OTS) were coated on sputtered Co-Cr-W-Ni alloy thin film and bulk Co-Cr-W-Ni alloy. OTS SAM coated alloy specimens were characterized using contact angle goniometry, Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), and atomic force microscopy (AFM). Contact angle analysis and FTIR suggested that ordered monolayers were coated on both sputtered and bulk alloy. XPS suggested the selective dissolution of cobalt from the alloy during the formation of OTS SAM. The bonding between the alloy and the OTS SAM was mainly attributed to Si-O-Cr and Si-O-W covalent bonds and a smaller contribution from Si-O-Co bonds. AFM images showed the distribution of islands of monolayers coated on the alloy. The height of monolayers in majority of the islands was closer to the theoretical length of fully extended OTS molecules oriented perpendicular to the surface. The stability of OTS SAM was investigated in tris-buffered saline at 37 °C for up to 7 days. Contact angle, FTIR, and XPS collectively confirmed that the monolayers remain ordered and bound to the alloy surface under this condition. This study shows that Co-Cr alloys can be surface modified using SAMs for potential biomedical applications.     

Integrity of functional self-assembled monolayers on hydrogen-terminated silicon-on-insulator wafers

Silicon-on-insulator (SOI) wafers are commonly used to design microelectronics, energy conversion, and sensing devices. Thin solid films on the surfaces of SOI wafers have been a subject of numerous studies. However, SOI wafers modified by self-assembled monolayers (SAMs) that can also be used as functional device platforms have been investigated to a much lesser extent. In the present work, tert-butoxycarbonyl (t-boc, (CH₃)₃-C-O-C(O)-)-protected 1-amino-10-undecene monolayers were covalently attached to a H-terminated SOI (1 0 0) surface. The modified wafers were characterized by X-ray photoelectron spectroscopy to confirm the stability of the SAM/Si interface and the integrity of the secondary amine in the SAM. The transmission electron microscopy investigation suggested that this t-boc-protected 1-amino-10-undecene SAM produces atomically flat interface with the 2 μm single crystalline silicon of the SOI wafer, that the SiO_x and both available Si/SiO_x interfaces are preserved, and that the organic monolayers are stable, with apparent thickness of 1.7 nm, which is consistent with the result of the density functional theory modeling of the molecular features within a SAM.