New Generation of Electrochemical Sensors Based on Multi-Walled Carbon Nanotubes

: Multi-walled carbon nanotubes (MWCNT) have provided unprecedented advances in the design of electrochemical sensors. They are composed by sp 2 carbon units oriented as multiple concentric tubes of rolled-up graphene, and present remarkable active surface area, chemical inertness, high strength, and low charge-transfer resistance in both aqueous and non-aqueous solutions. MWCNT are very versatile and have been boosting the development of a new generation of electrochemical sensors with application in medicine, pharmacology, food industry, forensic chemistry, and environmental ﬁelds. This work highlights the most important synthesis methods and relevant electrochemical properties of MWCNT for the construction of electrochemical sensors, and the numerous conﬁgurations and successful applications of these devices. Thousands of studies have been attesting to the exceptional electroanalytical performance of these devices, but there are still questions in MWCNT electrochemistry that deserve more investigation, aiming to provide new outlooks and advances in this ﬁeld. Additionally, MWCNT-based sensors should be further explored for real industrial applications including for on-line quality control. ﬁlm f GCE, amperometry, = 0.021 µ mol L − 1 stability to of commercial enzymatic biosensors. the positive synergistic effects that are observed when different nanomaterials are associated into the same the capacity versatility of the electrochemical sensors. also


Introduction
Several nanomaterials have been exploited for different purposes, but carbon nanotubes (CNT) and their composites emerged as the most exciting and versatile [1][2][3][4][5]. CNT are carbon allotropes with intermediate properties between graphite and fullerenes [6]. They are composed by sp 2 carbon units oriented as one (single-walled; SWCNT) or multiple concentric tubes (multi-walled; MWCNT) of rolled-up graphene, separated by ≈0.35 nm [4,[6][7][8]. Both nanostructures had positive impacts for the applied sciences, but the latter has gained more attention due to the higher yield and lower production cost per unit, thermochemical stability, and ability to maintain or improve its electrical properties when submitted to different functionalization processes [9]. A general MWCNT representation and the possible πand σ-covalent bonds established along the nanostructure are displayed in Figure 1. MWCNT also present remarkable active surface area, chemical inertness, high strength, and low charge-transfer resistance in both aqueous and non-aqueous solutions, making them suitable for numerous applications as nano-sized metallic and/or semiconducting components in transistors [10], energy storage [11], hybrid conductors [12], supercapacitors [13], thin-film coatings [2,5,6], field emitters [14], (bio)fuel cells [1][2][3]7], photovoltaic devices [1,3], electromagnetic shields [1,7], and electrochemical probes [1,2,[5][6][7]. MWCNT provide a series of functionalities to the electrochemical sensors, making them fantastic devices for analytical purposes. Electrochemical sensors are by definition, according the International Union of Pure and Applied Chemistry, "electrochemical devices that transform the effect of the electrochemical interaction analyte-electrode into a useful analytical signal" [15]; these effects may be promoted electrically or may be caused by a spontaneous interaction at the zero-current condition [15]. Thus, electrochemical sensors may be organized in four main subgroups: voltammetric, potentiometric, potentiometric solid electrolyte gas sensors, and chemically sensitized field effect transistor [15]. MWCNT associate high surface area (about 1600 m 2 g −1 ) and exceptional electrical conductivity (current densities as high as 10 9 A cm -2 ), which suit them to the miniaturization and portability of the systems [14].

Figure 2.
General layout of an (a) electrical arch-discharge (adapted by permission from Reference [23]), (b) laser ablation (adapted by permission from Reference [23]) and (c) chemical vapor deposition apparatus (adapted by permission from Reference [25]) used to synthesize multi-walled carbon nanotubes.
EAD is relatively less expensive, offers better yield quality, but involves high temperature for synthesis [21][22][23]. Triggering an electrical arc-discharge between two electrodes, a plasma composed by carbon and metallic catalysts vapor (iron, nickel, cobalt, yttrium, boron, cerium, among others) is  [23]), (b) laser ablation (adapted by permission from Reference [23]) and (c) chemical vapor deposition apparatus (adapted by permission from Reference [25]) used to synthesize multi-walled carbon nanotubes. EAD is relatively less expensive, offers better yield quality, but involves high temperature for synthesis [21][22][23]. Triggering an electrical arc-discharge between two electrodes, a plasma composed by carbon and metallic catalysts vapor (iron, nickel, cobalt, yttrium, boron, cerium, among others) is formed under a rare gas atmosphere (helium or argon) [21][22][23]. The vapor content is a consequence of the energy transferred to the catalysts-doped anode, causing its erosion, while MWCNT are formed as a hard and consistent deposit on the cathode [23]. The LA method also represents a promising alternative to synthesize high-quality carbon nanotubes at room temperature, although it demands high instrumental and operational costs [21][22][23]. In this technique, a pulsed or continuous laser beam is focused on catalyst-based graphite pellet, which is placed at the center of a quartz tube filled with an inert gas and kept at 1200 • C. The radiant energy is enough to sublimate the carbon substrate, and this vapor is swept by the gas flow towards the conical water-cooled copper collector. MWCNT deposits are formed in the majority on the same collector, but appreciable amounts are also found on the quartz tube walls and graphite pellet backside. Both EAD and LA also produce in parallel other carbon phases and all products contain metallic impurities from the catalysts [23]. The small fractions of remaining metals do not compromise the performance of the electrochemical devices, and there are several purification processes proposed in the literature and by some commercial companies to remove undesirable phases [22][23][24]. Nevertheless, these procedures are based on oxidation under strong acidic conditions, which may significantly affect the integrity of the nanostructures.
The synthesis performed by CVD overcomes the aforementioned difficulties, guarantees large-scale production and, for these reasons, it has become more widely used to obtain MWCNT [22][23][24][25]. This technique involves either a heterogeneous (if a solid substrate is used) or a homogeneous process (supposing that reactions takes place in the gas phase) [23,25]. The main synthetic route explores the catalytic decomposition of a carbon-containing source on substrates containing transition metals and their composites (Au, Ag, Cu, Cr, Co, Fe, Mn, Mo, Ni, Pt, Pd, SiO 2 , SiC, and ceramics), which enables the growth of aligned and dense arrays of nanotubes [23,24]. Alternative energy sources such as plasma and optical excitation can also be employed, giving the possibility to synthesize MWCNT at low-temperatures (450-1000 • C) compared to EAD and LA [23][24][25]. The CVD method also allows greater control of selectivity/type, homogeneity and size of the nanotubes (a few tens to hundreds of micrometers) in comparison to the others, although these characteristics are strongly dependent on the nature and structure of the catalysts, and the operational conditions of the synthesis. All these aspects affect the density and speed of electron transfer through the nanostructures that, in turn, directly impact the performance of the electrochemical sensors.

MWCNT Electrochemical Properties
MWCNT-based electrochemical sensors have remarkable advantages for analytical applications, such as enhanced detection sensitivity, low charge-transfer resistance (R tc ), electrocatalytic effect, and reduced fouling. It is also speculated that traces of metallic catalysts can add remarkable electroanalytical characteristics (positive synergistic effect) to the nanotubes, playing a supporting role for the development of the devices [6,7,[15][16][17][18][19][20][21][22][23][24][25][26]. There are still controversies about the electron transfer mechanism along the structural network: Some researchers believe that the sidewalls of the CNT are inert (i.e., the edge-plane-graphite-like open ends and defect sites control the phenomenon) [26][27][28], but some studies conducted with two-dimensional SWCNT have shown evidence of charge-transfer on the basal-plane [7,23,25]. Although the uncertainties about the fundamental contributions of MWCNT in electrochemical sensors, the number of MWCNT successful applications in highly complex systems does not stop growing [2,9,17,26].
Understanding the size-dependent features is an indispensable requirement to explore the fascinating properties of nanotubes in analytical tools and to move the electroanalytical researches forward. In this sense, it must be assumed that the electronic band structure of CNT as a hexagonal lattice, where each carbon atom is covalently bonded to three others via sp 2 molecular orbitals (the fourth valence electron in a p z -orbital hybridizes with all others to form a delocalized π-band) [27].
Like this, an even number of electrons are contained in the basic nanotube structure, whose conduction character will depend on its behavior as metal or semiconductor. This property can be assessed through the tight-binding electronic structure calculations, considering the translational vector (C h ) that connects two equivalent crystallographic sites (unitary vectors a 1 and a 2 ) and its coefficients (n 1 and n 2 ), according to the following Equation (1): so that a metallic behavior is observed when the result is an integer multiple of three, while a non-metallic/semiconductor profile is observed for other cases; a direct consequence of the different conduction band gaps [6]. The electron-transfer along the MWCNT are also influenced by the tube diameter (d t ), chiral angle (θ), and basic translation vector (T), as shown in Figure 3. Thus, the manner in which a graphene sheet is rolled determines the main electronic properties of the resulting nanotube [6,15,23,27].
Appl. Sci. 2018, 8, x 5 of 20 so that a metallic behavior is observed when the result is an integer multiple of three, while a nonmetallic/semiconductor profile is observed for other cases; a direct consequence of the different conduction band gaps [6]. The electron-transfer along the MWCNT are also influenced by the tube diameter (dt), chiral angle ( ), and basic translation vector (T), as shown in Figure 3. Thus, the manner in which a graphene sheet is rolled determines the main electronic properties of the resulting nanotube [6,15,23,27]. Observing the referred parameters on the tubulene lattice, dt values can be determined by Equations (2) and (3), which relate this variable with the circumferential length of the nanotube (L) and a hexagonal network constant (a): considering °= 1.42 Å for carbon nanotubes [6]: Regarding the , the expression that describes its magnitude can be written as follows: The vector T is used to specify the orthogonal orientation to the Ch axis of the nanotube, and its value can be calculated by: where ranging from d (when 1 − 2 and is not a multiple of 3d) to 3d (when 1 = 2 and is a multiple of 3d) for chiral tubulenes, considering d the greatest common divisor for the coefficients 1 and 2 [6,15,23,27]. Observing the referred parameters on the tubulene lattice, d t values can be determined by Equations (2) and (3), which relate this variable with the circumferential length of the nanotube (L) and a hexagonal network constant (a): considering a• = 1.42Å for carbon nanotubes [6]: Regarding the θ, the expression that describes its magnitude can be written as follows: The vector T is used to specify the orthogonal orientation to the C h axis of the nanotube, and its value can be calculated by: where d k ranging from d (when n 1 − n 2 and d k is not a multiple of 3d) to 3d (when n 1 = n 2 and d k is a multiple of 3d) for chiral tubulenes, considering d the greatest common divisor for the coefficients n 1 and n 2 [6,15,23,27]. In addition, a translation vector (τ) is also necessary to delimit the unitary cell of the nanotube. It can be represented by Equations (6)-(8), knowing that d r (d-3d) is the greatest common divisor of the respective coefficients. In summary, the limits of the 1D nanotube unit cell (plane segment OAB'BO) are demarcated by the vectors C h and T, whereas a 1 and a 2 define the area of the 2D graphene unit cell [6,27]. This sequence of equations is very important for the development of electrochemical sensors because it allows to determine the number of electric and phononic bands in the carbon nanostructures, i.e., the magnitude of the electrical conduction through the nanomaterial. The influence of the band structure on the nanotube conduction bands is defined in the context of the Fermi velocity (υ F ).
whereh is the reduced Planck constant, k F is the Fermi wavevector, and m represents the mass of the nanostructure under study. These values are not well-defined in cases where the Fermi surface is non-spherical, but it is estimated to be υ F ∼ 10 6 ms −1 for semiconductors or even higher for conductors [27]. This prevision corroborates the observation recorder in the density of states diagram for SWCNT with different chirality, since at the Fermi energy level (E F ) there is a finite electronic transition for metallic structures (first van Hove optical transition), but a zero-band gap for semiconductors (second van Hove optical transition). A general description of the band structures and density of states for metallic and semiconductor nanotube is presented in Figure 4. Thus, if we imagine that MWCNT are composed by several coaxial SWCNT, it might be expected that they are not strictly a 1D-type conductor [7,27]. Likewise, when the nanotubes have different chirality, the band structure undergoes alterations, and the resultant coupling between the electronic states at the E F and phonon modes may cause superconductivity [4,7,27]. Despite the similarity with metallic materials, the carrier density in carbon nanotubes (1D quantum wires) is much lower and novel physical electron-electron phenomena (such as spin-charge separation and suppression of the electron tunneling density of states) should be considered to justify the possible discrepancies, as predicted by Luttinger liquid theory [23,[26][27][28]. Some electrochemical studies showed a lower R tc for MWCNT containing structural defects and/or traces of metallic impurities (catalyst nanoparticles), so that the current flow occurs mainly through the outer most nanotube cylinder [26][27][28]. Only one of the concentric tubes needs to exhibit this behavior for the overall electronic properties to be essentially metallic-low quantum capacitance values [7,27]. It is believed that if d t is smaller than the elastic mean free path, the one-dimensional ballistic transport predominates, but the opposite characterizes a two-dimensional diffusive current flow [7,[26][27][28]. In addition, nanotubes with larger d t have a greater density of structural defects, besides facilitating their modification and functionalization, which are interesting requirements for the development of electroanalytical devices [2,[6][7][8][9]. The mean length of the nanotubes also has a significant effect on the electron-transfer rate; they are inversely proportional properties [22,24,27]. Vertically aligned structures also exhibit superior performance compared to those randomly immobilized [2,16,[25][26][27][28].
imagine that MWCNT are composed by several coaxial SWCNT, it might be expected that they are not strictly a 1D-type conductor [7,27]. Likewise, when the nanotubes have different chirality, the band structure undergoes alterations, and the resultant coupling between the electronic states at the EF and phonon modes may cause superconductivity [4,7,27]. Despite the similarity with metallic materials, the carrier density in carbon nanotubes (1D quantum wires) is much lower and novel physical electron-electron phenomena (such as spin-charge separation and suppression of the electron tunneling density of states) should be considered to justify the possible discrepancies, as predicted by Luttinger liquid theory [23,[26][27][28].  Some electrochemical studies showed a lower Rtc for MWCNT containing structural defects and/or traces of metallic impurities (catalyst nanoparticles), so that the current flow occurs mainly through the outer most nanotube cylinder [26][27][28]. Only one of the concentric tubes needs to exhibit this behavior for the overall electronic properties to be essentially metallic-low quantum capacitance values [7,27]. It is believed that if dt is smaller than the elastic mean free path, the one-dimensional ballistic transport predominates, but the opposite characterizes a two-dimensional diffusive current flow [7,[26][27][28]. In addition, nanotubes with larger dt have a greater density of structural defects, besides facilitating their modification and functionalization, which are interesting requirements for the development of electroanalytical devices [2,[6][7][8][9]. The mean length of the nanotubes also has a significant effect on the electron-transfer rate; they are inversely proportional properties [22,24,27]. Vertically aligned structures also exhibit superior performance compared to those randomly immobilized [2,16,[25][26][27][28].
Therefore, MWCNT provide special characteristics in electrochemical sensors both in aqueous and non-aqueous electrolytes (electrocatalytic properties, high and fast electron-transfer, signal amplification and stability, among other particularities observed in different prototypes) when compared with the conventional carbonaceous electrodes (such as glassy carbon, highly-oriented pyrolytic graphite, carbon microfiber, and boron-doped diamond) [26][27][28]. However, the true reasons and fundamental electrochemical properties responsible for the reported performances (both when nanotubes are used alone or in (nano)composites) are still not fully elucidated and demand further Therefore, MWCNT provide special characteristics in electrochemical sensors both in aqueous and non-aqueous electrolytes (electrocatalytic properties, high and fast electron-transfer, signal amplification and stability, among other particularities observed in different prototypes) when compared with the conventional carbonaceous electrodes (such as glassy carbon, highly-oriented pyrolytic graphite, carbon microfiber, and boron-doped diamond) [26][27][28]. However, the true reasons and fundamental electrochemical properties responsible for the reported performances (both when nanotubes are used alone or in (nano)composites) are still not fully elucidated and demand further research [2,4,6,9,17].

Overview of MWCNT Applications in Electrochemical Sensors
Different and exciting properties are obtained with macro-, micro-, and nano-structured carbon derivatives [4,17], but the implementation of the latter in the configuration of electrochemical sensors brought new opportunities to detect trace concentrations (µmol L -1 to pmol L -1 ) of numerous analytes (active principle and metabolites of drugs, personal care products, flame retardants, gases, surfactants, Appl. Sci. 2018, 8, 1925 8 of 18 industrial additives, polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), hormones, pesticides, among others) in complex matrices (natural waters, soils, foodstuffs, biological fluids, fuels, wastes, and even in living organisms) [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17]. This trend led to the development of the newest category of electroanalytical devices (third generation/way), i.e., nanotools for nanoscale analysis [2,16]. The basic mechanism for the operation of these tools is based on the conversion of redox processes, registered at the sensor/electrolyte interface, onto quantitative or semi-quantitative information. MWCNT had an important role to reach such advances, and this fact can be easily attested by the exceptional growth of scientific reports in this field [2][3][4][5][6]8,16,17]. MWCNT's large active surface area, facile electron-transfer with a variety of molecules, and their use in nano-composites (mixed with conductive polymers, nanoparticles and active biomolecules), as well as their high potential for device miniaturization, have been enhancing the sensitivity of current methodologies and providing new sensing opportunities [2,17,[19][20][21][22]. The multichannel electron ballistic transport and the presence of holes/defects along the structures, especially when functionalized with carboxyl groups and their derivatives, further enhance the previously mentioned properties [2,[4][5][6][7][8][9]. When converting a graphene sheet to a nanotube, one electron of the carbon atoms remains in p-orbitals, organized between valence (π) and conduction (π*) bands with very low bandgap (a partial σ-π hybridization), which favors a fast electron transfer. The density of states in MWCNT structure depends on diameter (≈2-100 nm), chirality/configuration (concentric, herringbone, and bamboo) and nature (semiconductor, conductor, and superconductor) [1,7,15]. In addition, the strength of C−C bonds is among the most robust and, when they are organized in the nanotube form, these interactions also provide flexibility and resistance to rupture, increasing the versatility and number of possible applications of these nanomaterials [4,7,16].
Electrochemical sensing of a wide variety of chemically, biologically, and environmentally important analytes was investigated on MWCNT (chemical purity ≥95%)-based solid electrodes, as shown in Table 1. Numerous traditional and modern pharmaceuticals, for instance, are electroactive on these sensors and relevant contributions have been reported, either in stationary or in flow-through systems. Mao et al. [29] fabricated an electrochemical sensor for metronidazole (antibacterial and anti-inflammatory compound employed for the treatment of protozoal diseases) based on MWCNT and chitosan (CTS)-nickel complex modified glassy carbon electrode (GCE) by the self-assembly technique. The polymer matrix facilitates the immobilization of the nanocomposite on the electrode support, while the association between nickel and carbon nanomaterials increases the current intensity of the electroanalytical signal. Using differential pulse voltammetry (DPV), Ni-CTS/MWCNT/GCE also showed electrocatalytic effect toward metronidazole reduction, making the method highly selective to its analysis in pharmaceutical and biological samples, with a low detection limit (LOD; 0.025 µmol L −1 ) and suitable reproducibility and stability. Holanda et al. [30] reported a simple methodology to quantify acetaminophen (a compound with analgesic and antipyretic action) in commercial formulations, using a GCE modified with gold nanoparticles (Au-NP), functionalized MWCNT (f -MWCNT) and cobalt phthalocyanine (Co-Pht) as working sensor. Using f -MWCNT, a lower system capacitance and better stability of the sensor was observed. The quantification of acetaminophen on Co-Pht/f -MWCNT/Au-NP/GCE was carried out by square-wave voltammetry (SWV) that, in turn, showed a sensitivity (LOD = 0.135 µmol L −1 ) and reproducibility similar to the standard ultraviolet-visible spectrophotometric procedure (at the wavelength of 257 nm). Montes et al. [31] studied the superior electrochemical activity of MWCNTs with shorter dimensions (diameter × length: 100-170 nm × 5-9 µm and 6-9 nm × 5 µm) towards the oxidation of propionic acid derivatives, which are drugs with analgesic, antipyretic, and anti-inflammatory properties. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetric (CV) data showed better electrocatalysis effect, R tc and signal amplification for the sensor constructed with the shorter diameter nanostructures due to the increased defect density revealed by Raman spectroscopic measurements. The developed sensor (MWCNT (diameter × length: 6-9 nm × 5 µm)/GCE) was adapted to a flow-injection analysis system (FIA) and provided a sensitivity and accuracy to determine ibuprofen (LOD = 1.9 µmol L −1 ) by amperometry comparable to that obtained by capillary electrophoresis. Pavinatto et al. [32] used a fluorine doped tin oxide (FTO) electrode, coated with nanostructured layer-by-layer films of CTS@MWCNT (Figure 5), as a sensor for 17-α-ethinylestradiol, that is a synthetic estrogen widely used as an oral contraceptive. The electron-transfer on the sensor reduced dramatically in the presence of the nanotubes, improving its electroanalytical performance and enabling assays at very low concentrations. Using the irreversible and adsorption-controlled oxidation process of the estrogen onto the three-bilayer CTS@MWCNT/FTO, it was possible to determine it by SWV (LOD = 0.09 µmol L −1 ) without significant effect of the most common interfering compounds present in human urine (urea, uric, and ascorbic acids, glucose, NaCl, KCl, and NH 4 Cl). Chen et al. [33] used a nanocomposite based on MWCNT and 1-octyl-3-methyl-imidazolium hexa-fluorophosphate ionic liquid (IL) to modify a GCE, to develop a chiral electrochemical sensor for the enantiomeric recognition of propranolol, using the linear sweep voltammetry (LSV) technique. The authors did not present a LOD but declared the success of the proposed method to evaluate the enantiomeric purity of reagents and wastewater treatment efficiency. Other promising MWCNT-based electrochemical devices were also reported to successfully analyze hydrochlorothiazide and triamterene [34], mangiferin and icariin [35], natamycin [36], omeprazole [37], gentamicin [38], and many other drugs [2,6,9,[17][18][19][20], attesting their electroanalytical performance and effectiveness.
Appl. Sci. 2018, 8, x 9 of 20 fluorophosphate ionic liquid (IL) to modify a GCE, to develop a chiral electrochemical sensor for the enantiomeric recognition of propranolol, using the linear sweep voltammetry (LSV) technique. The authors did not present a LOD but declared the success of the proposed method to evaluate the enantiomeric purity of reagents and wastewater treatment efficiency. Other promising MWCNTbased electrochemical devices were also reported to successfully analyze hydrochlorothiazide and triamterene [34], mangiferin and icariin [35], natamycin [36], omeprazole [37], gentamicin [38], and many other drugs [2,6,9,[17][18][19][20], attesting their electroanalytical performance and effectiveness. The growing trend of using miniaturized systems for the electroanalysis of bioactive molecules in living organisms through less invasive tests has positive repercussion and wide acceptance [2,17]. From this perspective, glucose sensors have been receiving special attention, since these analytic tools can be used as an effective way to diagnose diabetes, hypoglycemia, and other metabolic diseases in real time, using human blood and other secreted fluids as study matrix. Başkaya et al. [39] fabricated a non-enzymatic glucose sensor using a hybrid film (composed by f-MWCNT and highly monodisperse nickel nanoparticles) immobilized on GCE, which was able to determine this target in human blood serum samples by amperometry, with sensitivity (LOD = 0.021 μmol L −1 ) and stability similar to one of commercial enzymatic biosensors. The results also support the positive synergistic effects that are observed when different nanomaterials are associated into the same matrix, enhancing the capacity and versatility of the electrochemical sensors. Purine bases are also crucial biomolecules to the immune function and abnormal changes causes serious disorders, such as cancer, epilepsy, lupus erythematous, renal calculi, and cardiovascular alterations. Wang et al. [40] prepared a nanocomposite by mixing MWCNT and copper-nickel hybrid nanoparticles (Cu@Ni-NP), which was deposited onto a GCE surface, with the goal of monitoring adenine and guanine in ds-DNA from mice brain tissues. Using DPV under the optimized conditions, the Cu@Ni-NP/MWCNT/GCE sensor achieved a LOD of 0.17 μmol L −1 and 0.33 μmol L −1 for guanine and adenine, respectively, attesting to its potential to quantify purine bases in complex samples. Likewise, sensors for point-of-care detection of DNA damage in medical diagnostics are also on the rise. Li and Lee [41] investigated the The growing trend of using miniaturized systems for the electroanalysis of bioactive molecules in living organisms through less invasive tests has positive repercussion and wide acceptance [2,17]. From this perspective, glucose sensors have been receiving special attention, since these analytic tools can be used as an effective way to diagnose diabetes, hypoglycemia, and other metabolic diseases in real time, using human blood and other secreted fluids as study matrix. Başkaya et al. [39] fabricated a non-enzymatic glucose sensor using a hybrid film (composed by f -MWCNT and highly monodisperse nickel nanoparticles) immobilized on GCE, which was able to determine this target in human blood serum samples by amperometry, with sensitivity (LOD = 0.021 µmol L −1 ) and stability similar to one of commercial enzymatic biosensors. The results also support the positive synergistic effects that are observed when different nanomaterials are associated into the same matrix, enhancing the capacity and versatility of the electrochemical sensors. Purine bases are also crucial biomolecules to the immune function and abnormal changes causes serious disorders, such as cancer, epilepsy, lupus erythematous, renal calculi, and cardiovascular alterations. Wang et al. [40] prepared a nanocomposite by mixing MWCNT and copper-nickel hybrid nanoparticles (Cu@Ni-NP), which was deposited onto a GCE surface, with the goal of monitoring adenine and guanine in ds-DNA from mice brain tissues. Using DPV under the optimized conditions, the Cu@Ni-NP/MWCNT/GCE sensor achieved a LOD of 0.17 µmol L −1 and 0.33 µmol L −1 for guanine and adenine, respectively, attesting to its potential to quantify purine bases in complex samples. Likewise, sensors for point-of-care detection of DNA damage in medical diagnostics are also on the rise. Li and Lee [41] investigated the performance of f -MWCNT (functionalization with oxygen-containing groups to improve the immobilization and stability of the bioelement) as an additive in electrochemical DNA sequence differentiation ability. The electroanalytical signal achieved by DPV was highly selective and modulated between the probe (5 -GTG TTG TCT CCT AGG TTG GCT CTG-3 ; 24-base fraction of the p53 gene, used as a marker for breast cancer) and its complementary sequence (5 -CAG AGC CAA CCT AGG AGA CAA CAC-3 ). The detection capacity (LOD = 141.2 pmol L −1 ) was more than two orders better than the previously reported for graphene functionalized (f -Grf) based sensors, even in the presence of single (5 -CAG AGC CAA CCT CGG AGA CAA CAC-3 ) and all-base mismatched pairs (5 -ATA TCG ACC TTG GCC GAG ACG GTG-3 ). Great effort has also been directed towards the development of electrochemical devices capable of early detection of immune deficiency diseases, such as acquired immune deficiency syndrome (AIDS) caused by the HIV-retrovirus. Ma et al. [42] proposed a device composed by a GCE modified with MWCNT and molecularly imprinted polymers (MIPs) based on CTS for the determination of HIV-p24 protein in real human serum samples ( Figure 6). Using DPV and the developed sensor (HIV-p24/CTS/MWCNT/GCE), a low limit of detection (0.083 pg cm −3 ), good selectivity, repeatability, reproducibility, stability, and accuracy were reached for the proposed electroanalysis; validation was performed by comparative studies with enzyme-linked immunosorbent assay (ELISA). The outstanding ability of MWCNT-based electrochemical sensors to simulate redox reactions that occur in vivo has also been demonstrated with the electroanalysis of hemoglobin [43], amino acids [44], cholesterol [45], bilirubin [46], neurotransmitters, and related compounds [47][48][49], and even microorganisms [50], in mammalian fluid samples, achieving great results and contributing to the early diagnosis and treatment of several related diseases.
The occurrence of pesticides in foodstuffs, natural water, soils, among other matrices of environmental relevance, constitute a worldwide concern. Sipa et al. [51] associated square-wave adsorptive stripping voltammetry (SWAdSV) and a modified sensor (β-cyclodextrins/MWCNT/GCE) to determine trace concentrations of the pesticide dichlorophen. The excellent electroanalytical performance of the system, mainly in the presence of nanotubes (LOD = 4.4 × 10 −8 mol L −1 ), allowed to monitor this vermicide in river water (Pilica River, in Poland) with good selectivity and absence of natural and anthropogenic interfering compounds. Özcan and Gürbüz [52] developed a simple and sensitive sensor for the voltammetric determination of the herbicide clopyralid in urine, river water, sugar beet, wheat, and herbicide formulations (Phaeton ® and Lontrel™). This novel sensor was built by modification of a GCE with a nanocomposite containing acid-activated MWCNT (oxygen-functionalized nanostructures with very low R tc ) and fumed silica, which, when associated with DPV, attained a LOD of 0.8 nmol L −1 , confirming the sensitivity of the proposed method. Ghodsi and Rafati [53] reported a strategy to develop a voltammetric sensor for diazinon (an organophosphate insecticide) based on TiO 2 nanoparticles (TiO 2 -NP) and MWCNT nanocomposite assembled onto a GCE. Under optimum conditions, TiO 2 -NP@MWCNT/GCE reached a LOD of 3.0 nmol L −1 , using both CV and SWV, and this performance was explored for the electroanalysis of the pesticide in well and tap water. Other mentioned advantages were the fast response time, good repeatability, stability, besides the fast and inexpensive electrode modification. Sensitive electrochemical sensors that associate the unique properties of MWCNT and metallic nanoparticles were also proposed by Wei et al. [54] and Ertan et al. [55], aiming to electroanalysize trichlorfon insecticide (LOD = 4.0 × 10 −7 mol L −1 , using Nafion ® /TiO 2 -NP@MWCNT@carboxymethyl chitosan/GCE) and simazine herbicide (LOD = 2.0 × 10 −11 mol L −1 , using MIP/platinum nanoparticles@polyoxometalate@f -MWCNT/GCE), respectively, reinforcing the positive synergistic effect between these nanostructures for the improvement of the electroanalytical methodologies. linked immunosorbent assay (ELISA). The outstanding ability of MWCNT-based electrochemical sensors to simulate redox reactions that occur in vivo has also been demonstrated with the electroanalysis of hemoglobin [43], amino acids [44], cholesterol [45], bilirubin [46], neurotransmitters, and related compounds [47][48][49], and even microorganisms [50], in mammalian fluid samples, achieving great results and contributing to the early diagnosis and treatment of several related diseases. The occurrence of pesticides in foodstuffs, natural water, soils, among other matrices of environmental relevance, constitute a worldwide concern. Sipa et al. [51] associated square-wave adsorptive stripping voltammetry (SWAdSV) and a modified sensor (βcyclodextrins/MWCNT/GCE) to determine trace concentrations of the pesticide dichlorophen. The excellent electroanalytical performance of the system, mainly in the presence of nanotubes (LOD = 4.4 × 10 −8 mol L −1 ), allowed to monitor this vermicide in river water (Pilica River, in Poland) with good selectivity and absence of natural and anthropogenic interfering compounds. Ö zcan and Gürbüz [52] developed a simple and sensitive sensor for the voltammetric determination of the herbicide clopyralid in urine, river water, sugar beet, wheat, and herbicide formulations (Phaeton ® and Lontrel™). This novel sensor was built by modification of a GCE with a nanocomposite containing acid-activated MWCNT (oxygen-functionalized nanostructures with very low Rtc) and fumed silica, The wide active area and low R tc observed for MWCNT, and related (nano)composites, have been providing important advances on the single and simultaneous electroanalytical quantification of metallic cations (Cd 2+ [56], Pb 2+ [56], Mn 2+ [57], and Na + [58]) and anions (SO 3 2− [59], 60], and BrO 3 − [60]) in varied and complex matrices (wastewater [57], drinking water [56], and groundwater [58,59]). Trace concentrations of persistent organic pollutants (sunset yellow [61], tartrazine [62], and luteolin [62] dyes), as far as gases (chlorine [63], carbon dioxide [64], and methanol vapor [65]) and industrial by-products (glycerol [66], bisphenol A [67], hydrazine [68], and hydrogen peroxide [69]) are now being monitored with MWCNT-based sensors in a quick, reproducible, reliable, and cost-effective way when compared to the traditional analytical protocols, including those based on electrochemical devices from previous generations.

Final Remarks
MWCNT and their (nano)composites have been the origin for exciting and versatile sensing platforms due to their inherent advantages, namely large active surface area, electrocatalytic properties, high and fast electron-transfer, signal enlargement, reduction of the overpotential, chemical inertness, among others. Their incorporation into electrochemical sensors has been improving the electroanalytical performance of these devices and simultaneously widening their scope and range of applications. However, some limitations related to the high cost of production of high purity MWCNT have been restricting their large-scale and industrial utilization. Thus, improvements on the current synthetic routes and purification methods or/and development of novel low cost and effective techniques are clearly needed. In addition, comprehensive characterization of the toxicity of MWCNT is another requirement to expand their application to in vivo assays. These achievements will contribute to the construction of reliable and reproducible MWCNT-based electrochemical sensors enlarging sensors exploitation in real in vivo monitoring and in on-line manufacturing applications. Funding: Simone Morais is grateful for financial support from the European Union (FEDER funds through COMPETE) and National Funds (Fundação para a Ciência e Tecnologia-FCT) through projects UID/QUI/50006/2013 and AAC No. 02/SAICT/2017-project No. 029547 "CECs(Bio)Sensing-(Bio)sensors for assessment of contaminants of emerging concern in fishery commodities", by FCT/MEC with national funds and co-funded by FEDER. She also thanks financial support by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF): projects Norte-01-0145-FEDER-000011 and Norte-01-0145-FEDER-000024.