**4. Chemical sensing**

Before exploring the use of CNTs in chemical and electrochemical sensing it should be pointed out that the term sensor refers to a device that responds to a physical or chemical stimulus by producing a signal, usually an electrical one (Hillberg et al., 2005), while a biosensor is a sensor that uses biological selectivity to limit perception to the specific molecule of interest. A typical biosensor consists of two main components: the chemosensory materials (receptors) that can selectively bind target analytes and the efficient transducer that can transform the binding events into a readable signal output related to the analyte concentration in the sample (Eggins, 2002). The efficiency of chemosensors is largely dependent on the selectivity and sensitivity of the used sensory materials to a target species. In the traditional approaches a biological or biologically derived sensing element acting as a receptor is immobilized on the surface of a physical transducer to provide selective binding of analytes (Orellana & Moreno-Bondi, 2005; Jiang & Ju, 2007). As a sensing element, it is possible to use either biological macromolecules (e.g. antibodies, enzymes, receptors and ion channel proteins, nucleic acids, aptamers and peptide nucleic acids) or biological systems (e.g. ex vivo tissue, microorganisms, isolated whole cells and organelles). However, the small surface area and non-tunable surface properties of transducers greatly limit the efficiency of chemosensors, especially for the detection of ultratrace analytes. Recently, nanomaterials have found a wide range of applications as a material foundation of chemosensors, and have exhibited various degrees of success in the improvement of detection sensitivity and selectivity (Gao D. et al., 2007; Xie et al., 2006; Banholzer et al., 2008) due to their unique electrical, optical, catalytic or magnetic properties (Chen J.R. et al., 2004) and large surface-to-volume ratio (Xie et al., 2008).

#### **4.1 CNTs in chemical sensing**

Among nanomaterials, carbon surfaces represent very attractive materials for electrochemical studies, such as biosensor applications, due to their different allotropes (graphite, diamond and fullerenes/nanotubes). Carbon electrodes are well polarizable. However, their electrical conductivity strongly depends on the thermal treatment, microtexture, hybridization and content of heteroatoms. Additionally, the amphoteric character of carbon allows use of the rich electrochemical properties of this element from donor to acceptor state. Application of recently developed carbon materials include,

temperatures CVD grown CNTs exhibit a lower crystallinity than tubes produced using the

When considering the whole of CNTs applications, the solubilization of pristine CNTs in aqueous solvents remains an obstacle to realizing their potential, due to the rather hydrophobic character of the graphene sidewalls. To successfully disperse CNTs the dispersing medium should be capable of both wetting the hydrophobic tube surfaces and modifying the tube surfaces to decrease tube aggregation. Four basic approaches have been used to obtain a dispersion: surfactant-assisted dispersion (Ham et al., 2005; Islam et al., 2003; Moore et al., 2003; Yurekli et al., 2004; Vaisman et al., 2006), solvent dispersion (Fu & Sun, 2003; Ausman et al., 2000; Kim D.S. et al., 2005), functionalization of CNT sidewalls (Dyke & Tour 2004; Fernando et al., 2004; Peng et al., 2003), and biomolecular dispersion

Before exploring the use of CNTs in chemical and electrochemical sensing it should be pointed out that the term sensor refers to a device that responds to a physical or chemical stimulus by producing a signal, usually an electrical one (Hillberg et al., 2005), while a biosensor is a sensor that uses biological selectivity to limit perception to the specific molecule of interest. A typical biosensor consists of two main components: the chemosensory materials (receptors) that can selectively bind target analytes and the efficient transducer that can transform the binding events into a readable signal output related to the analyte concentration in the sample (Eggins, 2002). The efficiency of chemosensors is largely dependent on the selectivity and sensitivity of the used sensory materials to a target species. In the traditional approaches a biological or biologically derived sensing element acting as a receptor is immobilized on the surface of a physical transducer to provide selective binding of analytes (Orellana & Moreno-Bondi, 2005; Jiang & Ju, 2007). As a sensing element, it is possible to use either biological macromolecules (e.g. antibodies, enzymes, receptors and ion channel proteins, nucleic acids, aptamers and peptide nucleic acids) or biological systems (e.g. ex vivo tissue, microorganisms, isolated whole cells and organelles). However, the small surface area and non-tunable surface properties of transducers greatly limit the efficiency of chemosensors, especially for the detection of ultratrace analytes. Recently, nanomaterials have found a wide range of applications as a material foundation of chemosensors, and have exhibited various degrees of success in the improvement of detection sensitivity and selectivity (Gao D. et al., 2007; Xie et al., 2006; Banholzer et al., 2008) due to their unique electrical, optical, catalytic or magnetic properties (Chen J.R. et al.,

Among nanomaterials, carbon surfaces represent very attractive materials for electrochemical studies, such as biosensor applications, due to their different allotropes (graphite, diamond and fullerenes/nanotubes). Carbon electrodes are well polarizable. However, their electrical conductivity strongly depends on the thermal treatment, microtexture, hybridization and content of heteroatoms. Additionally, the amphoteric character of carbon allows use of the rich electrochemical properties of this element from donor to acceptor state. Application of recently developed carbon materials include,

two alternative methods mentioned above.

2004) and large surface-to-volume ratio (Xie et al., 2008).

**4.1 CNTs in chemical sensing** 

(Gigliotti et al., 2006).

**4. Chemical sensing** 

environment-friendly and energy-saving automotive parts, high-purity components in semiconductor manufacturing equipment for highly integrated chips, internal walls of vacuum vessels in the nuclear fusion reactors expected to be an energy source in the 21st century, and negative electrodes of lithium ion secondary batteries, of which demand is rapidly increasing with the popularization of mobile electronic instruments (Kazua, 1999). During the last years a great interest has been focused on the application of carbons (Flavel et al., 2009; Ma G.P. et al., 2009) as electrode materials because of their accessibility, easy processability and relatively low cost. They are chemically stable in different solutions (from strongly acidic to basic) and able to show performance in a wide range of temperatures. Already well-established chemical and physical methods of activation allow for the production of materials with a developed surface area and a controlled distribution of pores that determine the electrode/electrolyte interface for electrochemical applications.

The possibility of using the activated carbon without binding substances (e.g. fibrous fabrics or felts) gives an additional advantage from the construction point of view. Unique properties of ultra-microelectrodes (UMEs) make them very attractive in electroanalytical measurements which require high spatial and temporal resolution (Wigthman, 1988). At ultra-microelectrodes, at typical slow scan rates of voltammetry, signal to noise ratios (S/N) can increase up to one decade due to efficient mass transportation to the electrode resulting from edge effects. In addition, because of the small dimensions and low IR drop, fast scan voltammetric measurements (Garreau et al., 1990; Hsueh et al., 1993) and measurements in highly resistive media are possible at UMEs (Howell & Wightman, 1984) while high spatial resolution and minimal tissue damage (Musallam et al., 2007). can be achieved in vivo analysis. Microelectrodes are useful for measurements for changes in easily oxidized neurotransmitter concentrations in extracellular brain fluid (Kawagoe et al., 1993) as they provide a way to observe the rapid chemical changes associated with the release of neurotransmitters from neurons and their subsequent removal from the extracellular fluid (Atesa & Sarac, 2009).

The nanodimensions of CNTs guarantee a very large active surface area and are especially suited for the conception of miniaturized sensors. In addition to the high porosity and reactivity, this makes them ideal candidates for the storage of neutral species as well as electron donors, when used as electrodes in electrochemical reactions, for which they may also be called "nanoelectrodes" (Basabe-Desmonts et al., 2007; Daniel et al., 2007; Guo et al., 2004). Hence, it is not surprising that gas sensors made from individual nanotubes show good sensitivities at room temperature (J. Kong et al., 2001; Vasanth Kumar et al., 2010, 2011) in comparison to commercially available classical semiconductor sensors, which in general operate above 200°C. However, a necessary prerequisite is that the molecules to be detected must have a distinct electron donating or accepting ability, which is fulfilled, for example, by ammonia (NH3) as a donor and nitrogen dioxide (NO2) as an acceptor. The adsorption of these molecules on the nanotubes is associated with a partial charge transfer, which alters the charge-carrier concentration or, alternatively, the adsorbed molecules may affect the potential barriers present at the tube–electrode contacts. In any event, the resulting change in the electrical resistance of the nanotube is utilized as a sensor signal. However, for the detection of molecules that are only weakly adsorbed (e.g., carbon monoxide and hydrogen), the change in resistance is often too small. A possible method to overcome this drawback is accomplished by the modification of the nanotube sidewalls with nanoparticles made of a suitable metal. For instance, sensitive hydrogen sensors operating at room

Carbon Nanotubes – Imprinted Polymers: Hybrid Materials for Analytical Applications 189

(Zhao G.C. et al., 2003), as well as glucose oxidase (Guiseppi-Elie et al., 2003). It is noteworthy that in the latter case, the redox-active center is deeply embedded within the protein. In some cases, an oxidative pretreatment that introduces negatively charged surface groups on the CNTs was necessary to achieve high electron transfer rates. In a strategy to optimize the accessibility of the redox center, aligned CNT arrays have been fabricated using self-assembly techniques, followed by the covalent attachment of microperoxidase to the tube ends (Gooding et al., 2003). On this basis, various types of amperometric biosensors have been fabricated (Rubianes et al., 2003; Wang J. & Musameh, 2003; Wang J. et al., 2003). A glucose sensor, for example, is obtained by immobilizing glucose oxidase onto SWCNTs deposited on a glassy carbon surface (Davis et al., 2003). Hrapovic et al., 2004 combined platinum nanoparticles with carbon nanotubes and developed a new approach for the electrochemical detection of glucose (Aponte et al., 2006). Effective deposition of Pt nanoparticles on SWCNT was achieved by using a negatively charged polymer, Nafion, to dissolve and disperse SWCNT. Pt nanoparticles can easily be deposited on the Nafionmodified CNT due to charge interactions. Compared to the sensors containing only Pt or

CNT, the integration of Pt and CNT lead to improved sensitivity and detection limit.

redox potential, with cobalt acting as the mediator.

labels and a conventional glassy carbon transducer.

**4.2 MIPs in chemical sensing** 

Zhang and Gorski used carbon nanotubes and redox mediators dispersed into a polymeric matrix for NADH detection and found a reduction of the overpotential (by 0.3 V), a higher sensitivity, and faster response times (Zhang M. & Gorski, 2005). Wang J. & Musameh, 2004 reported on the use of carbon nanotubemodified glassy carbon electrodes in order to accelerate the electrooxidation of insulin, which resulted in a lowering of the required detection potential and a 14 nM detection limit as studied by flow injection analysis. Wang K. et al., 2005 proposed a potentiometric sensor for ascorbic acid on the basis of cobalt phthalocyanine nanoparticles directly coated on a glassy carbon electrode by drop coating. The authors explained the selective and rapid response to ascorbic acid on the basis of the

Kerman et al., 2004 also described the label-free electrochemical detection of DNA based on the direct attachment of adenine probes to the sidewall and end of functionalized MWCNTs. The MWCNTs were attached onto the carbon paste electrode surface modified with thymine probes by hybridization between adenine and thymine. The combination of sidewall and end functionalization of MWNT showed enhancement of the guanine oxidation signal in the

The attachment of numerous enzymes on a single carbon nanotube label provided for enormous signal enhancement. Wang J. et al., 2004 reported on a novel method to dramatically amplify enzyme-based DNA sensing by using carbon nanotubes. In the new sensing scheme, the DNA duplex acted as connectors between microbeads and CNTs. The CNT loaded with enzymes linked to the magnetic microbeads through the interaction of complementary oligonucleotides. Without the recognition event, the CNT cannot attach to the particles and is removed during the magnetic separation. The CNTs function as carriers for enzyme tags, and the enzyme reaction products were also accumulated in the CNTs. The coupling of enzyme-tagged CNTs provided a better detection limit than single-enzyme

In the sensors and biosensors technology, often there is the overriding complication that it is unusual, in "real" samples, for there to be a single species present. More commonly an

direct measurement compared to the ones from only end-modified MWCNT.

temperature can be obtained via the deposition of palladium nanoparticles either by direct evaporation (Kong J. et al., 2000) or through electrodeposition (Geng et al., 2004; Teles et al., 2008). Electrodeposition offers the specific advantage of site-selectivity, since the metal decoration is restricted to the current-carrying tubes, so that the remaining substrate surface is unaffected. An example is a semiconducting SWCNT with electrodeposited Pd particles upon exposure to hydrogen. The operation mechanism is largely analogous to that of a palladium gate field-effect transistor realized within classical silicon technology (Lundstroem et al., 1975). Specifically, molecular hydrogen is split on the surface of a Pd particle into atomic hydrogen, which diffuses to the Pd/SWCNT interface. At this interface, a dipole layer is formed, which acts like a microscopic gate electrode that locally changes the charge-carrier concentration. It should be mentioned that the recovery of this type of roomtemperature-operated hydrogen sensor requires a supply of oxygen to remove the hydrogen atoms in the form of water (Balasubramanian & Burghard, 2005). Carbon nanotubes exhibit high electron transfer rates for different redox couples in various media (Balasubramanian et al., 2004) which has stimulated an increasing amount of research into CNT based amperometric sensors for the detection of specific analytes in solution. The length scales of CNTs are similar to that of typical biological molecules, which gives CNTs an edge over other materials in functioning as effective electrodes in bioelectrochemical sensing (Guiseppi-Elie et al., 2002). In particular, their high aspect ratio and their diameter in the nanometer range make CNTs particularly well suited for direct electrochemical communication with the redox site of a protein, without requiring any mediator. When properly arranged, a nano- tube should have the capability to act as a 1D channel that guides electrons towards the redox center. These materials can be used to preconcentrate analytes and for the magnetic separation and molecular identification of biomolecules, and organic and inorganic species (AguilarArteaga et al., 2010).

The application of CNTs as absorption and/or transducing materials was shown in several different works. Recently, a critical review dealing with the adsorption mechanism of analytes of different nature on carbon nanotubes was published (Cruz et al., 2010; Fontanals et al., 2007; See et al., 2010; Petrovic et al., 2010; Zeng J. et al., 2009; Augusto et al., 2010; Lucena et al., 2011;). The authors stated that this procedure cannot be explained by a single mechanism but through a combination of mechanisms. Indeed, the introduction of functional groups on the surface can facilitate the selective interaction with a given family of compounds. However, this fact has not yet been exploited for the analysis of urine samples. MWCNTs are proposed as a sorbent in an off-line SPE method for the determination of antidepressants using LC and UV detection (Bakker & Qin, 2006). MWCNTs have been applied as adsorbents for preconcentration of sulfonylurea herbicides (Lemos et al., 2008; Zhou Q. et al., 2006a, 2007), bisphenol A, 4-n-nonylphenol, 4-tert-octylphenol (Cai et al., 2003a), dichlorodiphenyltrichloroethane and its metabolites (Zhou Q. et al., 2006b); atrazine, simazine (Zhou Q. et al., 2006c), and trihalomethanes (Lu et al., 2005) in environmental water samples, barbiturates determination in pork (Zhao H. et al., 2007), for simultaneous determination of 10 sulfonamides in eggs and pork (Fang et al., 2006), for trapping volatile organic compounds in a purge-and trap GC system (Li Q. et al., 2004), to extract diethylphthalate, di-n-propyl-phthalate, di-iso-butyl-phthalate, and di-cyclohexylphthalate from aqueous solutions (Cai et al., 2003b).

Direct electron transfer has been achieved with various types of CNT electrodes for cytochrome c (Wang J. et al., 2002), horseradish peroxidase (Zhao Y. et al., 2002), myoglobin

temperature can be obtained via the deposition of palladium nanoparticles either by direct evaporation (Kong J. et al., 2000) or through electrodeposition (Geng et al., 2004; Teles et al., 2008). Electrodeposition offers the specific advantage of site-selectivity, since the metal decoration is restricted to the current-carrying tubes, so that the remaining substrate surface is unaffected. An example is a semiconducting SWCNT with electrodeposited Pd particles upon exposure to hydrogen. The operation mechanism is largely analogous to that of a palladium gate field-effect transistor realized within classical silicon technology (Lundstroem et al., 1975). Specifically, molecular hydrogen is split on the surface of a Pd particle into atomic hydrogen, which diffuses to the Pd/SWCNT interface. At this interface, a dipole layer is formed, which acts like a microscopic gate electrode that locally changes the charge-carrier concentration. It should be mentioned that the recovery of this type of roomtemperature-operated hydrogen sensor requires a supply of oxygen to remove the hydrogen atoms in the form of water (Balasubramanian & Burghard, 2005). Carbon nanotubes exhibit high electron transfer rates for different redox couples in various media (Balasubramanian et al., 2004) which has stimulated an increasing amount of research into CNT based amperometric sensors for the detection of specific analytes in solution. The length scales of CNTs are similar to that of typical biological molecules, which gives CNTs an edge over other materials in functioning as effective electrodes in bioelectrochemical sensing (Guiseppi-Elie et al., 2002). In particular, their high aspect ratio and their diameter in the nanometer range make CNTs particularly well suited for direct electrochemical communication with the redox site of a protein, without requiring any mediator. When properly arranged, a nano- tube should have the capability to act as a 1D channel that guides electrons towards the redox center. These materials can be used to preconcentrate analytes and for the magnetic separation and molecular identification of biomolecules, and

The application of CNTs as absorption and/or transducing materials was shown in several different works. Recently, a critical review dealing with the adsorption mechanism of analytes of different nature on carbon nanotubes was published (Cruz et al., 2010; Fontanals et al., 2007; See et al., 2010; Petrovic et al., 2010; Zeng J. et al., 2009; Augusto et al., 2010; Lucena et al., 2011;). The authors stated that this procedure cannot be explained by a single mechanism but through a combination of mechanisms. Indeed, the introduction of functional groups on the surface can facilitate the selective interaction with a given family of compounds. However, this fact has not yet been exploited for the analysis of urine samples. MWCNTs are proposed as a sorbent in an off-line SPE method for the determination of antidepressants using LC and UV detection (Bakker & Qin, 2006). MWCNTs have been applied as adsorbents for preconcentration of sulfonylurea herbicides (Lemos et al., 2008; Zhou Q. et al., 2006a, 2007), bisphenol A, 4-n-nonylphenol, 4-tert-octylphenol (Cai et al., 2003a), dichlorodiphenyltrichloroethane and its metabolites (Zhou Q. et al., 2006b); atrazine, simazine (Zhou Q. et al., 2006c), and trihalomethanes (Lu et al., 2005) in environmental water samples, barbiturates determination in pork (Zhao H. et al., 2007), for simultaneous determination of 10 sulfonamides in eggs and pork (Fang et al., 2006), for trapping volatile organic compounds in a purge-and trap GC system (Li Q. et al., 2004), to extract diethylphthalate, di-n-propyl-phthalate, di-iso-butyl-phthalate, and di-cyclohexylphthalate

Direct electron transfer has been achieved with various types of CNT electrodes for cytochrome c (Wang J. et al., 2002), horseradish peroxidase (Zhao Y. et al., 2002), myoglobin

organic and inorganic species (AguilarArteaga et al., 2010).

from aqueous solutions (Cai et al., 2003b).

(Zhao G.C. et al., 2003), as well as glucose oxidase (Guiseppi-Elie et al., 2003). It is noteworthy that in the latter case, the redox-active center is deeply embedded within the protein. In some cases, an oxidative pretreatment that introduces negatively charged surface groups on the CNTs was necessary to achieve high electron transfer rates. In a strategy to optimize the accessibility of the redox center, aligned CNT arrays have been fabricated using self-assembly techniques, followed by the covalent attachment of microperoxidase to the tube ends (Gooding et al., 2003). On this basis, various types of amperometric biosensors have been fabricated (Rubianes et al., 2003; Wang J. & Musameh, 2003; Wang J. et al., 2003). A glucose sensor, for example, is obtained by immobilizing glucose oxidase onto SWCNTs deposited on a glassy carbon surface (Davis et al., 2003). Hrapovic et al., 2004 combined platinum nanoparticles with carbon nanotubes and developed a new approach for the electrochemical detection of glucose (Aponte et al., 2006). Effective deposition of Pt nanoparticles on SWCNT was achieved by using a negatively charged polymer, Nafion, to dissolve and disperse SWCNT. Pt nanoparticles can easily be deposited on the Nafionmodified CNT due to charge interactions. Compared to the sensors containing only Pt or CNT, the integration of Pt and CNT lead to improved sensitivity and detection limit.

Zhang and Gorski used carbon nanotubes and redox mediators dispersed into a polymeric matrix for NADH detection and found a reduction of the overpotential (by 0.3 V), a higher sensitivity, and faster response times (Zhang M. & Gorski, 2005). Wang J. & Musameh, 2004 reported on the use of carbon nanotubemodified glassy carbon electrodes in order to accelerate the electrooxidation of insulin, which resulted in a lowering of the required detection potential and a 14 nM detection limit as studied by flow injection analysis. Wang K. et al., 2005 proposed a potentiometric sensor for ascorbic acid on the basis of cobalt phthalocyanine nanoparticles directly coated on a glassy carbon electrode by drop coating. The authors explained the selective and rapid response to ascorbic acid on the basis of the redox potential, with cobalt acting as the mediator.

Kerman et al., 2004 also described the label-free electrochemical detection of DNA based on the direct attachment of adenine probes to the sidewall and end of functionalized MWCNTs. The MWCNTs were attached onto the carbon paste electrode surface modified with thymine probes by hybridization between adenine and thymine. The combination of sidewall and end functionalization of MWNT showed enhancement of the guanine oxidation signal in the direct measurement compared to the ones from only end-modified MWCNT.

The attachment of numerous enzymes on a single carbon nanotube label provided for enormous signal enhancement. Wang J. et al., 2004 reported on a novel method to dramatically amplify enzyme-based DNA sensing by using carbon nanotubes. In the new sensing scheme, the DNA duplex acted as connectors between microbeads and CNTs. The CNT loaded with enzymes linked to the magnetic microbeads through the interaction of complementary oligonucleotides. Without the recognition event, the CNT cannot attach to the particles and is removed during the magnetic separation. The CNTs function as carriers for enzyme tags, and the enzyme reaction products were also accumulated in the CNTs. The coupling of enzyme-tagged CNTs provided a better detection limit than single-enzyme labels and a conventional glassy carbon transducer.

#### **4.2 MIPs in chemical sensing**

In the sensors and biosensors technology, often there is the overriding complication that it is unusual, in "real" samples, for there to be a single species present. More commonly an

Carbon Nanotubes – Imprinted Polymers: Hybrid Materials for Analytical Applications 191

furthermore, they are produced via complex protocols with a high cost and require specific handling conditions because of their poor stability (Whitcombe et al., 2000; Wulff, 2002;

During the past ten years, the literatures on the development of MIP-based sensors, in particularly electrochemical (Riskin et al., 2008;) and optical (McDonagh et al., 2008; Basabe-Desmonts et al., 2007; Li J. et al., 2007a; Feng et al., 2008) sensors, have been dramatically growing (Nilsson et al., 2007; Ramanavicius et al., 2006). It was found that the manufacture of composites consisting of molecularly imprinted conducting polymers results in obtaining materials that exhibit both predetermined selective molecular recognition and electrical conductivity (Deore et al, 2000). This type of materials, mostly frequented based on overoxidized polypyrrole, is of special interest for use in the field of sensor technology

MIP-based electrochemical sensors were first reported in the early 1990s by Mosbach's group (Andersson et al., 1990), and to date, remarkable progress in MIP-based electrochemical sensors have been achieved in the use and the performance of conductometric/potentiometric MIP nanomaterials (Zhou Y.X. et al., 2003), which were used to detect many different analytes (Augisto et al., 2010) such as barbituric acid (Mirsky et al., 1999), amino acid derivatives (Panasyuk et al., 1999), morphine (Kriz et al., 1995), atrazine (Kim **Y.** et al., 2007), benzyltriphenylphosphonium chloride (Kriz & Mosbach, 1995), thiophenol (Kröger et al., 1999), glutamic acid (Ouyang et al., 2007), folic acid (Prasad et al., 2010a; Prasad et al., 2010c), tolazoline (Zhang Z. et al., 2010a), tryptophan (Prasad et al., 2010d; Kong Y. et al., 2010), clindamycin (Zhang J. et al., 2010), 2,4-dichlorophenoxy acetic acid (Xie et al, 2010), histamine (Bongaers et al., 2010), caffeine (Alizadeh et al., 2010; Vinjamuri et al., 2008), theophylline (Kan et al., 2010) uracil and 5-fluorouracil (Prasad et al., 2009), salicylic acid (Kang et al., 2009), uric acid (Patel et al., 2009), resveratrol (Xiang & Li, 2009), hydroquinone (Kan et al., 2009; Kan et al., 2008a), bisphenol (Kolarz & Jakubiak,

Despite the application of MIPs as sensor matrices or separation materials, they suffer from basic limitations associated with the limited concentration of imprinted sites, and the bulk volume of the polymer matrices that requires long diffusion paths of the imprinted host molecules. These limitations lead to inefficient sensing or separation processes (Shi et al., 2007). In order to increase the efficiency of MISPE, a method is increasing the surface area of polymer. Higher surface area can contain more functional groups, thus the interaction between polymer and bioactive compounds will be increased accordingly (Tian et al., 2011). A useful approach is the fabrication of robust multilayer structures by the photocrosslinking of the layers (Sun et al., 1998, 2000). This process transformed the electrostatic interlayer stabilizing interactions into covalent bonds. This achievement, together with the stated limitations of MIPs, suggested that the surface imprinting of LbL nanostructured

**5. CNTs-MIPs composites: innovative materials for analytical determinations**  Among the different applications of CNTs in nanotechnology, polymer composites, consisting of additives and polymer matrices, including thermoplastics, thermosets and elastomers, are considered to be an important group of relatively inexpensive materials for many engineering applications (Ma et al., 2010; Yang et al., 2007). The technology implications are significant to

films might be a viable technique to fabricate effective MIP matrices.

Haupt & Mosbach, 2000; Ye & Haupt, 2004).

2008), and dopamine (Kan et al., 2008b).

(Shiigi et al., 2003).

analyte of interest is accompanied by a number of different species, all present at different concentrations and all adding to the complexity of the analytical problem.

All over the world, billions of dollars are spent annually on chemical/biological detections related to medical diagnosis, environmental monitoring, public security and food safety because lab analysis using expensive equipment is usually cumbersome and timeconsuming. Therefore, there has been a pressing societal need for the development of chemo/biosensors for the detection of various analytes in solution and atmosphere, which are both less expensive and simpler to construct and operate. Although considerable progress was made in the past several decades, the chemo/biosensor field remains underdeveloped and at a low level of commercialization because of the lack of alternative strategies and multidisciplinary approaches (Guan et al., 2008).

The standard approach to the analytical analysis of complex matrices is the separation of the different components. Typically, therefore, before a sensor can be used to perceive and quantify one component in a mixed solution, the various components of the complex mixture must be separated, usually by a chromatographic process, so that some form of non-selective sensor, e.g. UV absorbance measurements, can be used to detect and quantify each individual component. In order to improve the performance of chemical sensors, an improvement of their selectivity is required, so that a particular chemical species can be detected and assayed without the need for a possibly lengthy separation stage. In this direction, a technological approach is the development of the biosensor (Updike & Hicks, 1967).

Molecular imprinting is one of the most efficient strategies that offer a synthetic route to artificial recognition systems by a template polymerization technique (Ye & Mosbach, 2001; Spivak, 2005; Zhang H.Q. et al., 2006). While most research in this direction is targeted to the design of chromatographic stationary phases, their use in electrochemical sensors is expanding for electroactive analytes. In convincing work, the group of Mandler explored sol-gel polymers imprinted with the organophosphate pesticide parathion and performed gasphase and liquid-phase partitioning experiments as well as cyclic voltammetric studies (Marx et al., 2004). Imprinted films showed a more than 10-fold increased equilibrium binding over nonimprinted polymers and discriminated well against a range of other structurally similar organophosphates.

To date molecularly imprinted polymers have been successfully used with most types of transduction platforms and a range of methods have been used to bring about close integration of the platform with the polymer (Adhikari & Majumdar 2004). By using MIP it is possible to overcome the commonly founded limitations in the traditional biosensor approach (Orellana & Moreno-Bondi, 2005; Jiang & Ju, 2007), based on the use of biological macromolecules (e.g. antibodies, enzymes, receptors and ion channel proteins, nucleic acids, aptamers and peptide nucleic acids) or biological systems (e.g. ex vivo tissue, microorganisms, isolated whole cells and organelles) as sensing element. In the last years, indeed, although biological receptors have specific molecular affinity and have been widely used in diagnostic bioassays and chemo/biosensors, great affords have been made in synthesizing artificial recognition receptors to overcome the limit in the efficiency of chemosensors, especially for the detection of ultratrace analytes due to the small surface area and non-tunable surface properties of transducers. Another limitation of traditional biosensors is that often natural receptors for many detected analytes do not exist;

analyte of interest is accompanied by a number of different species, all present at different

All over the world, billions of dollars are spent annually on chemical/biological detections related to medical diagnosis, environmental monitoring, public security and food safety because lab analysis using expensive equipment is usually cumbersome and timeconsuming. Therefore, there has been a pressing societal need for the development of chemo/biosensors for the detection of various analytes in solution and atmosphere, which are both less expensive and simpler to construct and operate. Although considerable progress was made in the past several decades, the chemo/biosensor field remains underdeveloped and at a low level of commercialization because of the lack of alternative

The standard approach to the analytical analysis of complex matrices is the separation of the different components. Typically, therefore, before a sensor can be used to perceive and quantify one component in a mixed solution, the various components of the complex mixture must be separated, usually by a chromatographic process, so that some form of non-selective sensor, e.g. UV absorbance measurements, can be used to detect and quantify each individual component. In order to improve the performance of chemical sensors, an improvement of their selectivity is required, so that a particular chemical species can be detected and assayed without the need for a possibly lengthy separation stage. In this direction, a technological approach is the development of the biosensor (Updike & Hicks,

Molecular imprinting is one of the most efficient strategies that offer a synthetic route to artificial recognition systems by a template polymerization technique (Ye & Mosbach, 2001; Spivak, 2005; Zhang H.Q. et al., 2006). While most research in this direction is targeted to the design of chromatographic stationary phases, their use in electrochemical sensors is expanding for electroactive analytes. In convincing work, the group of Mandler explored sol-gel polymers imprinted with the organophosphate pesticide parathion and performed gasphase and liquid-phase partitioning experiments as well as cyclic voltammetric studies (Marx et al., 2004). Imprinted films showed a more than 10-fold increased equilibrium binding over nonimprinted polymers and discriminated well against a range of other

To date molecularly imprinted polymers have been successfully used with most types of transduction platforms and a range of methods have been used to bring about close integration of the platform with the polymer (Adhikari & Majumdar 2004). By using MIP it is possible to overcome the commonly founded limitations in the traditional biosensor approach (Orellana & Moreno-Bondi, 2005; Jiang & Ju, 2007), based on the use of biological macromolecules (e.g. antibodies, enzymes, receptors and ion channel proteins, nucleic acids, aptamers and peptide nucleic acids) or biological systems (e.g. ex vivo tissue, microorganisms, isolated whole cells and organelles) as sensing element. In the last years, indeed, although biological receptors have specific molecular affinity and have been widely used in diagnostic bioassays and chemo/biosensors, great affords have been made in synthesizing artificial recognition receptors to overcome the limit in the efficiency of chemosensors, especially for the detection of ultratrace analytes due to the small surface area and non-tunable surface properties of transducers. Another limitation of traditional biosensors is that often natural receptors for many detected analytes do not exist;

concentrations and all adding to the complexity of the analytical problem.

strategies and multidisciplinary approaches (Guan et al., 2008).

1967).

structurally similar organophosphates.

furthermore, they are produced via complex protocols with a high cost and require specific handling conditions because of their poor stability (Whitcombe et al., 2000; Wulff, 2002; Haupt & Mosbach, 2000; Ye & Haupt, 2004).

During the past ten years, the literatures on the development of MIP-based sensors, in particularly electrochemical (Riskin et al., 2008;) and optical (McDonagh et al., 2008; Basabe-Desmonts et al., 2007; Li J. et al., 2007a; Feng et al., 2008) sensors, have been dramatically growing (Nilsson et al., 2007; Ramanavicius et al., 2006). It was found that the manufacture of composites consisting of molecularly imprinted conducting polymers results in obtaining materials that exhibit both predetermined selective molecular recognition and electrical conductivity (Deore et al, 2000). This type of materials, mostly frequented based on overoxidized polypyrrole, is of special interest for use in the field of sensor technology (Shiigi et al., 2003).

MIP-based electrochemical sensors were first reported in the early 1990s by Mosbach's group (Andersson et al., 1990), and to date, remarkable progress in MIP-based electrochemical sensors have been achieved in the use and the performance of conductometric/potentiometric MIP nanomaterials (Zhou Y.X. et al., 2003), which were used to detect many different analytes (Augisto et al., 2010) such as barbituric acid (Mirsky et al., 1999), amino acid derivatives (Panasyuk et al., 1999), morphine (Kriz et al., 1995), atrazine (Kim **Y.** et al., 2007), benzyltriphenylphosphonium chloride (Kriz & Mosbach, 1995), thiophenol (Kröger et al., 1999), glutamic acid (Ouyang et al., 2007), folic acid (Prasad et al., 2010a; Prasad et al., 2010c), tolazoline (Zhang Z. et al., 2010a), tryptophan (Prasad et al., 2010d; Kong Y. et al., 2010), clindamycin (Zhang J. et al., 2010), 2,4-dichlorophenoxy acetic acid (Xie et al, 2010), histamine (Bongaers et al., 2010), caffeine (Alizadeh et al., 2010; Vinjamuri et al., 2008), theophylline (Kan et al., 2010) uracil and 5-fluorouracil (Prasad et al., 2009), salicylic acid (Kang et al., 2009), uric acid (Patel et al., 2009), resveratrol (Xiang & Li, 2009), hydroquinone (Kan et al., 2009; Kan et al., 2008a), bisphenol (Kolarz & Jakubiak, 2008), and dopamine (Kan et al., 2008b).

Despite the application of MIPs as sensor matrices or separation materials, they suffer from basic limitations associated with the limited concentration of imprinted sites, and the bulk volume of the polymer matrices that requires long diffusion paths of the imprinted host molecules. These limitations lead to inefficient sensing or separation processes (Shi et al., 2007). In order to increase the efficiency of MISPE, a method is increasing the surface area of polymer. Higher surface area can contain more functional groups, thus the interaction between polymer and bioactive compounds will be increased accordingly (Tian et al., 2011). A useful approach is the fabrication of robust multilayer structures by the photocrosslinking of the layers (Sun et al., 1998, 2000). This process transformed the electrostatic interlayer stabilizing interactions into covalent bonds. This achievement, together with the stated limitations of MIPs, suggested that the surface imprinting of LbL nanostructured films might be a viable technique to fabricate effective MIP matrices.
