T
he creation of organized thin films (OTFs) of organic compounds has developed into an increasingly important research area at the frontier of analytical chemism ( 1 4 This importance stems from the critical role of liquid-solid and gas-solid interfaces in a host of emerging transduction mechanisms and from the relatively high degree of structural definition affordedby OTFs. In the case of liquid-solid interfaces, modification of an electrode surface can transform a comparatively nonselective electron-transfer process into one with an enhanced specificity based on the ionic charge (5,S), hydrophobicity or hydrophilicity (71, size ( 2 , 8-11), and identity (12) of an electroactive species. In the case of gas-solid interfaces, OTFs provide opportunities for creating interfaces that have well-defined composition, thickness, spatial orientation, and packing density and can be used to gain control of specificity and the extent of interactions between an analyte and a modi-
Chuan-Jian Zhong and Marc D. Porter Iowa State University 0003-2700950367-709A509000 ::1995 Amercan Cnemca Socelr
Organized thin films can be used to create interfacesfor manipulating reactions between analytes and modified surfaces
tural attributes of OTFs that can be used to clarify the issues related to transduction mechanisms that are important to analytical chemistry and to designing interfacial architectures with advanced functions. We highlight a few of the many uses of OTFs, beginning with fundamental studies of interfacial processes, followed by emerging applications to chemical analysis. Reviews of applications of OTFs in other areas provide additional fundamental and technological information (14).
Preparative methods The underlying phenomenon that led to fied surface. These same attributes impart the creation of LB films was discovered many decades before the phenomenon of OTFs with many of the characteristics of model interfaces that can be used to ex- SA films (2).The first scientific experiments with LB films examined the calmplore the fundamental issues related to the predictive design of interfaces with tar- ing effect of an oil dispersed on water and were reported more than 200 years ago get pelformance specifications. by Benjamin Franklin. It was Langmuir, In this Report we present an overview however, who spearheaded the early sysof the development and application of OTFs to analytical chemistry We will dis- tematic investigations of organic films at the air-water interface. Blodgett subsecuss the origins and preparative methodquently developed techniques for the ologies of Langmuir-Blodgett (LB) and transfer of these films to solid surfaces. spontaneously adsorbed (SA) films, the two most used routes toward the creation These pioneering studies focused primarily on the preparation and transfer of of OTFs. We will also examine the strucAnalytical Chemistry, December I , 1995 709 A
densely packed structures of long-chain substrate combination, which takes advanfatty acids (e.g., barium stearate and cadtage of the high preferential affinity of mium arachidate). Advances since then gold for sulfur-based ligands, was trighave expanded the scope of transfer gered largely by the report of Nuzzo and modes and organizational formats and ad- Allara on the formation of functionalized dressed issues related to compression disulfides at gold substrates (2). and transfer mechanisms, the role of the As depicted in Figure lb, the formation substrate, the construction of mixed- of SA films is initialized by the strong affincomponent films, and film stability. ity of the head group of the adsorbate for The fabrication of LB films (2)relies on the substrate. For example, the chemithe formation and transfer of a single layer sorption of long-chain carboxylic acids at of amphiphilic molecules spread at the metal oxides proceeds through the formaair-water interface, the surface area of tion of the corresponding metal carboxywhich is slowly shrunk to yield a film with late salt (2). On the other hand, silanes a defined molecular packing density. Both (e.g., RSii?,&SiI,or R,SiX, where R is vertical and horizontal lifting techniques an alkyl chain and X is a chloro or alkoxy can then be used to transfer the preorgagroup) are tethered to a hydroxylated surnized layer onto a smooth substrate. The face by a S i 4 bond and can polymerize vertical transfer procedure, shown in Figinto 2-D cross-linkednetworks using RSii, ure la, is used to prepare both hydropho precursors ( I , 2). Sulfuriontaining combic and hydrophilic OTFs. The horizontal pounds (thiols, disulfides, and sulfides) transfer method, in which the substrate chemisorb at gold as the corresponding "touches" the top of a preorganized layer, thiolate. In these situations, cleavage of is used primarily for the construction of S-H, S-S, and S-C bonds is central to the hydrophilic OTFs. In both cases, multi- formation process (14). layer structures are formed by repetiIn all cases, the packing of the OTF retion of the transfer process. sults from the cohesive interactions b e The origins of SAfilms can be traced tween the alkyl chains and is iduenced by largely to the classic wettability studies of chain length, end group, solvent, immerZisman (2).Today, there are three gension time, substrate morphology, temperaeral preparative strategies, all of which ture, and several other factors (15).The rely on the immersion of an appropriate vast interest in applying OTFs has been substrate into a dilute solution of an adsor- prompted by the ease of preparing interbate precursor. The fust strategy is based faces with well-defined composition, on the chemisorption of long-chaincarbox- thickness, and spatial orientation in comylic acids at metals with native oxides parison, for example, to the morphological (e&, copper, aluminum, silver, and chroand compositionalheterogeneity at the mium) (2).These systems constitute the surface of a polymeric film. platforms used in the noted wettahility The LB and SA approaches to the constudies. The second strategy uses orstruction of OTFs have relative strengths ganosilanebased monolayers linked to and weaknesses that should be considhydroxylated surfaces. The advantages of ered within the context of the proposed this technique were first recognized by application. The LB technique provides Sagiv as a route to the constructionof oleo- precise control of surface concentration phobic monolayers with a controllable and can be used to modify most smooth molecular organization (2).This approach surfaces. Preparative flexibility, however, has recently been extended to the formais limited to structures derived from amtion of multilayer structures by Mallouk phiphilic molecules. Durability may also ( 1 ) by using metal phosphonates a s be an issue when structural stability is bridges between layers. dominated by the cohesive interactions The third strategy, which has gained a between neighboring chains. strong foothold in many research areas (2, The SA preparation method is more 3) besides analytical chemistry, is based straightfomard it yields a surface strucon organosulfur-derived monolayers ture that, through changes in the end chemisorbed at gold (2, 13, 14), a few group, can be used to modify the compoother metals (3),and semiconductors.The sition of surfaces regardless of size and immense interest in this adsorbateshape. The types of samples address710 A
Analytical Chemistry, December 1, 1995
Figure 1. Idealized depictions of film formation. (a) Vertical mode LE film transfer of a pre-
compressed amphiphilic layer of molecules at an air-water interface to a solid substrate. (b) Formation of an SA monolayer at a substrate from a solution containing adsorbate
precursors.
able are nevertheless defined by the specificity of the interactions between the head group and the substrate. In addition, the ability to control the surface compo sition by co-assembly depositions is more empirical in nature because of nonideal thermodynamic correlationsbetween surface and solution composition (16). In both types of preparations,the existence of phase-segregateddomains and the importance of molecular cooperativity remain significant but unresolved issues. Models and applications
Electroanalytical chemistry was one of the first areas to take advantage of the structural definition of OTFs, which have been used as models for probing fundamental issues of heterogeneous electron transfer ( I 7-27) and electrical doublelayer t h e ones (13,19,20)and as platforms for creating selective bamers to electron trans fer (9-12). There are three distinct but potentially overlapping pathways for electron transfer
of a solution-based redox species at insuterfaces. In one series of studies, colating OTFs: permeation through the film, assembled monolayers were fabricated discharge at structural defects, and elecfrom a mixture of ferrocenyl- and methyltron tunneling. Because defects (e.g., pin- terminated alkanethiols (22).Variations in holes, adsorbate vacancies, and gauche the immediate environment around the kinks) represent weaknesses in harrier ferrocenylmoiety, through changes in the properties that may degrade perforchain length of the methyl-terminated mance, many early explorations focused component,markedly altered the thermoon assessing the structural integrity of dynamics of the redox process. Thus the OTFs (13,21).It was found, for examapparent reduction potential of a "buried" ple, that the increase in the length of the ferrocenylmoiety was shifted nearly 0.3 polymethylene chain in alkanethiolate V positive of that for the same moiety exmonolayers at gold effectively increased posed to the electrolytic solution. This is the "dielectric thickness'' of the OTF (13). attributed to the increased difficulty in In one instance, this system appeared to the generation and solvation of the posibe effective as a bamer in the delineation tive charge created through the oxidation of key parameters in Marcus theory (e.g., of the ferrocenyl group in a strongly hydrocarbon-like environment. tunneling coefficient and reorganization energy) using solution-based redox couIn a related study (2@,an electrochemples (18).In contrast, the packing limitaical quartz crystal microbalance investigation imposed by the larger relative size of tion probed the importance of the solvathe doxy head group relative to the alkyl tion and identity of the counterions on the chain yields a more disordered chain structure that proved less effective as a barrier for such evaluations (21). Another approach to a critical assessment of issues relevant to heterogeneous electron-transfer kinetics involves the use of multilayer systems (1) and the use of redox species pendant to thiolatebased monolayers (17,23-27). This strategy pins down the movement of the redox species, precluding possible complications from the electron transfer of a solution-based species at defects. Further processing by dilution of the redox sites within an inert methyl-terminated chain thermodynamicsof the electrochemical structure eliminates contributionsfrom transformation of monolayers formed from lateral electron-transfer processes. By a viologen species functionalized on one comparison, studies of OTFs at the airend with an alkyl chain and on the other water interface have proven effective in with a thiol-containing appendage. Systemdelineating phenomena involved in lateral atic changes in the length of the chains at electron-transfer processes (22).These both ends of the viologen species revealed and related strategies could senre as viathat the smaller the separation distance ble frameworks for probing the role of between the electrolytic solution and rethrough-space or through-bond mechadox site, the easier the solvation. The nisms and the importance of donorlarger solvation stabilized the dicationic form of the redox species, driving the acceptor separation distance, temperature, electrolyte, and solvent. oneelectron generation of the viologen Systems based on OTFs have also been monocation to more negative applied p e used as starting points for unraveling the tentials. details of how the microscopic environApproaches for architectures that have ment surrounding a redox site influpathways for vectorial electron and charge ences reactivity (3,23-27).m e goal is to conduction constitute another area in understand the predictive translation of which the structural features of OTFs are reactivity from the 3-D nature of gases and valuable (28-30).At issue are whether liquids to the lower dimensionality of inan anisotropic orientation of charge or
Electroanalytical chemistry was one of thefirst areas to take advantage of OTFs.
electron propagation pathways can be imparted to the surface structure and whether the resulting materials can function as wires or other components in molecular-based electronicdevices. Some of the more notable pathways have used LB or SA films with pendant charge-transfer complexes, such as tetrathiafulvalene and tetracyanoquinodimethane,metallophthalocyanines and metalloporphyrins (2,28),and rigid rod oligoimides (29). In most of these cases, as well as in efforts to fabricate 2-D networks of conducting polymers (e.g., polypyrrole [301), n-r interactions play an important role in the "stacking" of structures with directed conduction pathways. Opportunities for advancing the design of electrocatalysts by OTFs have also been explored. Single- and multilayer films from an amphiphilic derivative of a Ni(1I)containing cyclam were constructed by L5 methods at a glassy carbon electrode for the electrocatalytic reduction of CO, (31).The potency of the solution form of the catalyst was successfully translated to the surface-bound species. Similarly, various Cool) porphyrins were derivatized with alkanethiol appendages and anchored at gold by SA methods to probe both the stability and orientational aspects (32)of the efficiency for the tweelectron catalytic reduction of dioxygen to hydrogen peroxide. These findings will guide the synthetic design and formation of improved catalysts by OTFs of cofacial porphyrins in which the four-electron reduction of dioxygen has been demonstrated using adsorbed layers on carbon electrodes. Approaches to gain control over the specificity of reactions have also been pursued to enhance chemical discrimination in electrochemically based transduction mechanisms. Strategiesrange from control based on surface hydrophobicity to the exploitation of molecular recognition phenomena. Design issues center on the incorporation of a discrete set of physical and chemical properties into the interfacial structure while minimizing the consequences of defects. The interfacial properties of OTFs are dominated by the chemical identity of the end group (1,Z). Thus methyl-terminated OTFs are strongly hydrophobic, and carboxylic acid-terminated OTFs are strongly hydrophilic. These properties can
Analytical Chemistry, December 1, 1995 711 A
be used to manipulate the selectivity of rent represents an amplified analytical sig- voltammettic response of cytochrome c at electroanalytical measurements. In one ad- nal. uncoated gold is irreversible. The poor r e aptation, the selectivity of an amperometStructuralderivatizations using OTFs versihility is attributed to an unfolding of ric detector for LC was modified using have also had an impact in protein electre the protein upon adsorption at uncoated methyl-teminated alkanethiolatesat gold chemistry (W,33,34).One of the earligold and the improved reversibility to the (7). Because the hydrophobicity of this sys est ventures in using SA films was the enadsorption of cytochrome c in its native tem increases with the length of the alkyl hancement of electron-transfer rates for state at this and other compositionalvat+ chain, the preferential permeation of hydm cytochrome c at the interface formed by ants of %-modified electrodes (29. phobic analytes such as chlorpromazine the chemisorption of bis(4-pyridyl)disulStudies of other forms of interfacial reand promethazinethrough the longer chain fide at gold (33).By comparison,the activity have used the structural definistructuresprovided a basis for disnimination against hydrophilic anaiytes such as ferrocyanide and hydrogen peroxide. Concepts using electrostatics constiMicroporous tute a complementary route to electroanalytical selectiviy by OTFs (5. 6). For instance, the charge generated by the ioniza%Capture antibod tion of the carboxylic acid terminus of alkanethiolates at gold can be used to differentiate between ascorbic acid and dopamine (6). At physiological pH, dopamine is positively charged, whereas ascor) bic acid, which is present at much higher concentrations in neurological fluids, is negatively charged. Thus electrostatics diminish the elech-ooxidation of ascorbic acid and enhance that of dopamine. This approach is similar to that used with electrodes coated with much thicker films of the anionic polymer Nafion (3).However, the use of a single molecular layer as the surface modifier should improve r e sponse times and detection limits, enhancing performancefor in vivo applications. The complexation of analytes at OTFs '-I with chelating end groups is yet another route for improving electroanalytical selectivity. In one particularly intriguingexample (9,an indirect signal amplification process was used to enhance detection. 0 C This process derives from the creation of access channels to the electrode surface that result from the complexation of an analyte such as CaOI) with the end groups at lipid multilayer membranes (e.g., dide decyl phosphate) prepared by LB methods. These channels appear to form because of a combined complexationelectrostatic-induced conformational change in the structure of the membrane. Channel formation, and hence the presence of an analyte, is detected indi- Figure 2. i 4 c achemioal enzyme imunoassay. rectly by the voltammettic response of a (a)The capture antihuman chorionic gonadotropin (hCG) antibody immobilized by the redox species such as ferrocyanide. When derivatization of an SA-formedmonolayer of thioctic acid on a goid coating supported on a microporous membrane. (b) The capturing process of the immobilized antibody toward a protein the turnover of the redox species is P (hCG) in the sample in the presence of an enzyme-labeled conjugate antibody E. (c)The enzygreater than the amount of analyte bound matic reaction upon substrate diffusionfrom the back side of the membrane through pores and to the lipid membrane, the observed curelectrochemical detection 01 the resulting product. aminophenol. (Adapted from Reference38.) ~
U
712 A Analytical Chemistry, December 1, 1995
tion of OTFs. Many of these investigations have worked toward assessing interfacial acid-base reactivity, with the goal of addressing issues related to more complicated processes. Although it is clear that immobilization can induce large changes in acid-base reactivity (e.g., decreases in effective acid dissociation constants as large as lo3), the relative contributions of interfacial dielectric effects and intermolecular interactions have yet to be untangled. Efforts that model the doublelayer structure (19.20), coupled with systematic correlationsbetween structure and r e activity (23-271, promise to place such issues on firmer experimental and theoretical grounds. Tbe development of transductionpathways for optical, piezoelectric. and other forms of chemical sensors have taken several creative pathways using OTFs ( I ) . For example, combining the molecular information that can he obtained from Raman spectroscopy with the unique interfacial properties of OTFs poses a particularly attractive avenue of exploration. To this end, a partition-based interface was created to detect aromatic compounds by surface-enhanced Raman spectroscopy (SERS) (35).This approach used a SAformed monolayer of octadecanethiolate at a roughened silver surface, the resulting structure of which reversibly extracted aromatic organic compounds (e.g., 0-, m-, and pxylene) from aqueous solutions. The response of the sensor was internally calibrated against a vibrational mode of the alkanethiolate monolayer, a scheme that begins to address a long-standing barrier to Raman spectroscopic measurements in sensor applications.The eventual viability of this approach,which should have a clear impact on environmental monitoring needs, is strongly tied to the successful development of field-deployablehardware. Tbe surfaces of piezoelectric and other novel forms of sensor devices have been modified with OTFs (1).As in the electrochemical studies, differences in hydrophobicity-hydrophilicity, donor-acceptor, and complexation properties have been manipulated through changes in the end group to affect differences in the sorption of gaseous analytes ( 1 . 3 6 ) .Although the development of interfacial structures with a high degree of specificity remains an obstacle, approaches based on the design
of sensor arrays and multivariant data analysis are tractable pathways toward implementation ( I ) .
Advanced applications
Interest in improving specificity has driven the design of OTFs with more complex architectures, leading to new analysis concepts and variations of existing schemes. OTFs provide a platform from which an enormous number of schemes can be devised to exploit a complex superposition of chemical and physical interactionsfor affecting selectivity. In part, advances have focused on protocols for the synthesis of OTFs by adding sulfur- and silane-containing appendages to target moieties and by subsequent derivatization of the resulting layer. In the latter case, the carboxylic acid terminus of a long-chain alkanethiolate at
function” interfacial materials. In an early study, an amperometric sensor for glucose was constructed through the coimmobilization of the enzyme glucose oxidase (GOx) and a redox mediator 2-aminoethylferrocene (An. The linkages to gold were formed through a SA layer from 4-aminothiophenolthat was suhse quently activated with gluteraldehydefor cross-linking with the amine functional groups on both GOx and AF.Thus the two key components for an amperometric assay of glucose were afhed to the electrode surface. Equations 1-3 summarize the cooperative reaction pathway. The current for the regeneration of the oxidized form of AF (Equation 3) serYes as the analytical signal (39). GOx(oxidized) t glucose + GOx(reduced) + gluconolactone
(1)
GOx(reduced) + AF(oxidized) + GOx(oxidized) + AF(reduced) (2)
OTFs are emerging as an important factor in the miniaturization of the analytical laboratory. gold can be transformed to its acid chloride by treatment with thionyl chloride (37).This reaction is more effective when performed in a dry gaseous environment and reduces the possibility that the starting material will reform. Derivatizations have also taken advantage of the many coupling agents (e.g.. l-ethylW(3-dimethylaminopropy1)carhodiimide 1381) used in biochemical labeling. Each of these processes can be used to build a more complex interfacial structure through ester and amide linkages. The relative ease of subsequent derivatization is a key factor that has led to the immense interest in SA-based OTFs. Armed with these synthetic tools, researchers have created a variety of increasingly complex transducer interfaces. In the detection of biomolecules, efforts have often entailed the fabrication of “dual
AF(reduced)
+ AF(oxidized) t e- (3)
In another particularly intriguing scheme, an integrated structure based on the stepwise coupling of a capture antibody to a thioctic acid-derivatized, micro porous gold membrane was used to d e velop an enzyme immunoassay for p r o teins (Figure 2) (38).The concept exploits the binding of an analyte protein P that was previously incubated with an enzymelabeled conjugate antibody E to the capture antibody.Adding the enzyme substrate then generates an electroactive product by the immobilized enzyme that is amperometrically detected at the gold membrane. The performance of this system was competitive with that of existing methods and, more importantly, could be applied to assays of whole blood without loss of performance. By using a related pathway, SAfilms of a small synthetic peptide modified with a long alkanethiol appendage were tested as interfaces for the specific recognition of prcteins (40).Detection by surface plasmon resonance spectroscopy revealed both the high specxcity and the high affinity of an antibody-antigen reaction, but with a more preferential reversibility. The possible advantagesof the recognition properties found in solution-based guest-host chemistry has attracted the attention of several research groups. Routes
Anal!(fical Chemistrv, December 1, 1995 713 A
using both SA and LB methods for the organization of immobilized supramolecular molecules such as cyclodextrins (4, 8) and recesorcin[4]-arenes (36)have been explored. Two key features inherent in such architectures are that the "receptor" site must be oriented to facilitate accessibility by the analyte, which is required for efficientbinding, and the layer must be effectivelydevoid of defects, which is critical to minimize interference signals from processes that may occur at structural d e fects. Recognition by receptor sites provides opportunitiesfor devising detection strate gies for a wide range of analytes. Ionic selectivity has been achieved by the construction of a two
