Electrochemical sensor method for food quality evaluation

Electrochemical sensor method for food quality evaluation

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Neelam Yadav*,†, Annu Mishra*,†, Jagriti Narang*,† *MD University, Rohtak, India, †Amity Institute of Nanotechnology, Amity University, Noida, India

31.1

Introduction

Food security is a major global challenge because of the occurrence of hazardous chemical and biological toxins in food, which constitutes a threat to food supply and community well-being. World Health Organization (WHO) has stated that the foodborne diseases primarily damage the financial status of developing countries. Worldwide it has been reported, about 1500 diarrheal episodes occurred annually; among them, approximately 75% cases were caused by microbial contaminated food, and these were responsible for the deaths of nearly 3 million people in developing countries [1]. The US Centers for Disease Control and Prevention have reported that approximately 48 million US people become susceptible to foodborne diseases, of whom 128,000 people are hospitalized and 3000 die annually due to consumption of contaminated food [2]. Every year millions of lives are affected globally by outbreaks of foodborne pathogens that remain a major challenge to public health providers [2]. Foodborne pathogens can enter the body through contaminated water or contaminated and undercooked food. Hence, it is important to distinguish the presence of pathogens in food and water before it enters the body [1, 2]. Such organisms mainly include Acinetobacter spp., Bacillus subtilis, B. cereus, Campylobacter jejuni, Citrobacter koseri, C. freundii, Clostridium difficile, C. perfringens, Enterobacter sakazakii, E. cloacae, Escherichia coli O157:H7, Klebsiella oxytoca, K. pneumoniae, Listeria monocytogenes, Salmonella enteritidis, Salmonella typhimurium, Shigella sonnei, Staphylococcus aureus, Vibrio cholera, and Yersinia pestis. The major requirement for detection is in the areas of public health, the water and food industry and pharmaceutical industry [1]. Diagnosis and consequent treatment of infectious diseases, along with the detection of biohazards in the environment, is important for the classification of such microbes. Hence, problems related to foodborne diseases have been resolved by improving detection methods, which should be sensitive, selective, and offer quick response time [2]. Various techniques have been evolved to detect foodborne pathogens. The effort to improve the methods of detection has been a continuous process. The detection methods have been classified into conventional culture-based, immunological-based,

Evaluation Technologies for Food Quality. https://doi.org/10.1016/B978-0-12-814217-2.00031-7 © 2019 Elsevier Inc. All rights reserved.

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nucleic-based, and biosensor-based methods, each with their own principle, procedure, advantages, and limitations.

31.2

Conventional culture-based methods

Principle: Conventional culture testing methods depend on particular media to identify and isolate viable bacterial cells in food. These methods are very delicate, economical, and provide microbial information qualitatively as well as quantitatively present in the food sample. Advantages: Culture-based methods are the oldest methods for detecting microorganisms, even the pathogenic strains. These methods provide a confirmed result regarding the presence of a particular pathogen and the success rate is found to be high [3]. Limitations: The biggest drawback of culture-based methods is that they are time consuming, as they depend on the potential of the microbes to increase their population in varying nutrient media, such as pre-enrichment media, selective enrichment media, and selective plating. It must be noted that these entire media take up to 18–24 h to give the exact result, indicating the slow turnaround time. These traditional techniques employed for identification of pathogens causing foodborne diseases are slow because they require preparation of culture media, plate inoculation, and colony counting [4]. Example: One of the best-known examples which shows high success rate and also shows that the method is highly cost-effective. However, the process is time consuming and can produce pseudo positive results.

31.3

Immunological-based methods

Principle: Immunological techniques comprise of experimental procedures to investigate the immune system and to generate or employ immunological reagents as experimental devices. The widely used immune techniques involve the production and use of immunoglobulins for the determination of particular proteins in human samples. Antibody purity shows a significant role in success of the immunoassays [5]. Advantages: Developed immunoassays are easy to perform, provide fast result, and are less time consuming than culture-based methods. Hence, Enzyme linked immunosorbent assay (ELISA) has recently been a widely used immunoassay [5]. Limitations: These immunological assays are complicated, expansive, require sensitive reagents and are not suitable [5]. Examples: l

Tang et al. have developed a voltammetric sandwich immunosensor for detection of SEB in real food samples, which was found to have a better relationship with ELISA. The saturation kinetics of enzymes renders a tiny vibrant range for amperometric/voltammetric biosensors based on horseradish peroxidase-nanosilica-doped multiwalled carbon nanotubes (MWCNT) for amplified signal [6].

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Nucleic-based methods

Principle: These methods recognize DNA or RNA sequences of specific organisms isolated from the microorganism. Nucleic acid-based methods are explicit and fragile, and are employed for detection of various microorganisms. Various types of polymerase chain reaction (PCR) have emerged, which are given names according to the changed protocol of the original PCR [7]. Advantages: In general, the major advantages of PCR are that the process is rapid and sensitive. It is faster than the culture-based methods and immunoassays. The possible use of nucleic acid-based testing to the clinical laboratory is less time taken during diagnosis, high output, and exact and consistent results. PCR has now reached new heights, where the amplified product can be obtained in minutes, and distinguishing between the strains has become much easier as multiple primer pairs are used. PCR technique has developed as a very promising method of detection of the genes in pathogens; however, nucleic acid-based tests are qualitative, but existing quantification methods are limited and, hence, increase the risk for various infections such as HIV, cytomegalovirus, and human T-cell lymphotropic virus. However, due to their specificity, these methods are suitable for diagnosis and for monitoring response to treatment [8]. Limitations: During nucleic acid extraction and cell lysis there are more chances of cross-contamination and, hence, failure of reactions because of the presence of impeding substances or competing DNA from nonspecific cells. This can produce nonreliable results and reduces the value of PCR as a consistent strategy strategy [9]. Examples: l

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Isonhood et al. examined L. monocytogenes via PCR in mayonnaise-based RTE salads and obtained 105 CFU/g limit of detection for chicken, 104 CFU/g for macaroni, and 103 CFU/g for potato and seafood salads [10]. Lee et al. detected E. coli O157:H7, B. cereus, V. parahaemolyticus, Salmonella spp., L. monocytogenes, and S. aureus in Korean RTE food items such as chicken, steamed pork hocks, oysters, fresh tomato juice, sushi, and lettuce through an mPCR assay simultaneously. These pathogens were detected at 104 CFU/mL or greater in homogenized food samples [11]. Yang et al. performed a LAMP assay for the determination of enterotoxigenic E. coli from raw milk. By using this assay, the pathogen was detected at 547 CFU/mL of the pathogen. Foodborne bacterial pathogens have also been detected by modifying the LAMP assay; e.g., reverse transcription LAMP, complex LAMP, and in situ LAMP [12].

31.5

Recent technology development

31.5.1 Biosensor Basic Principle and Procedure: Biosensors are analytical devices that are comprised of recognition elements like DNA, RNA, enzymes, whole cells, and antibodies, coupled with transducing elements that measure physical activity, such as temperature, humidity, acceleration, distance, etc., after the alteration in chemical flow

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produced due to interaction with the analyte, and convert it into electrical current [13]. Specific analytes can be recognized by finding a molecular species to create a binding with the target pathogen for sensing. Various receptors have been used for augmenting the measuring efficiency of biosensors [14]. These biosensors are simple and costeffective and, hence, could be used in remote areas, where they would significantly enhance the quality of food and its safety by controlling exposure to microbes. Nanotechnology-based goods have offered a broad range of material candidates that increases portability, consistency, selectivity, sensitivity, and analytical performance of the sensors. The surface of the electrode was cleaned, dried to remove the contaminants present at the electrode surface, and nanomaterials were deposited onto the cleaned electrode. Then, biological recognition elements such as DNA, Aptamer, or Antibodies were immobilized onto the modified electrode and further analytes were added onto the biological recognition element modified electrode. Further electrochemical studies were recorded using the working electrode to confirm the different phases of electrode. Further different optimization studies, applicability, selectivity, and reproducibility tests were performed to check the bioassay for foodborne pathogen detection [15]. A biosensor is composed of two vital components: a bioreceptor and a transducer. The bioreceptor recognizes specific analytes, which can be as follows [13]: 1. Biological material: These are enzymes, antibodies, nucleic acids, and cell receptors. 2. Biologically derived material: These are aptamers and recombinant antibodies. 3. Biomimetic material: These include imprinted polymers and synthetic catalysts.

The transducer is involved in converting the biological interactions into a quantifiable analytical signal (Fig. 31.1). Advantages: Biosensing techniques have offered significant alternatives to avoid the drawbacks of traditional methods. They are rapid, portable, simple, and they do not require sample preenrichment (contrasting to conventional culture-based, nucleic-acid based and immunological-based methods which require sample preenrichment for concentrating the pathogens before detection). Recently, electrochemical biosensors have been used for the examination of foodborne pathogens [12].

31.5.2 Electrochemical-based biosensor A sensing electrode interacts with the analyte and, after interaction, the change in potential and current are observed; this is known as an electrochemical biosensor [7]. Electrochemical biosensors exhibit unique analytical characteristics compared to other transducing systems, like the possibility to function in cloudy media, better sensitivity, and capability of miniaturization (Scheme 31.1). Electrochemical biosensors have been classified depending on the type of sensing elements used, such as potentiometric and amperometric. The equipment used for electrochemical analysis is simple and cost-effective as compared to other analytical techniques [16]. However, these electrochemical biosensors possess some drawbacks similar to related biosensors, such as immobilizing of the bio-recognition component, devoid of denaturation or arbitrary direction. Therefore, most of the biosensors use

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Fig. 31.1 Schematic representation summarizing all electrochemical biosensors for the detection of foodborne pathogens.

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Scheme 31.1 Schematic representation of electrochemical biosensor for detection of specific DNA.

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nano-enabled platforms, as they exhibit unique substrates that assist for immobilization of the bio-recognition element through interactions like thiols, disulfides, amines, salines, or acids [7]. This chapter explains the progress made in the fabrication and development of electrochemical-based biosensors to evaluate food safety that can be achieved by NP-based detection frame; working factors and application for measuring contaminated food; challenges for practical employment; and discusses prospects for research directions.

31.5.2.1 Recent application progress in different types of foods l

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Wu et al. prepared an electrochemical sensor by immobilization of single-stranded DNA segments on gold electrodes via mercaptan of the DNA bases through N-hydroxysulfosuccinimide (NHS), and N-(3-dimethylamion) propyl-N0 -ethyl carbodiimide hydrochloride (EDC) as activator. Parameters like concentration of precise DNA and hybridizing conditions or the determination of sensitivity of the electrochemical assay have also been studied [17]. Wang et al. constructed a biosensor based on single-stranded DNA (ssDNA) probe functionalized aluminum anodized oxide (AAO) nanopore membranes or E. coli O157: H7 DNA detection [18]. Sun et al. fabricated an electrochemical DNA biosensor based on a dendritic nanogold and electrochemically reduced functionalized graphene carbon ionic liquid electrode for the detection of Listeria monocytogenes. The linear concentration range was 1.0  1012 to 1.0  106 mol/L with 2.9  1013 mol/L (3σ) LOD. This amperometric DNA sensor was selective and can distinguish one-base and three-base mismatched ssDNA sequences [19]. Bifulco et al. designed an electrochemical marker-free genosensor for scrutinizing inlA genes in Listeria strains. Single-stranded DNA was immobilized covalently at the surface of a gold electrode, hybridizes with complementary DNA sequences, and forms doublestranded DNA. Formation of this double stranded DNA reduced the current peaks of a differential potential voltammogram (DPV). This genosensor has the ability to differentiate entire DNA samples of L. monocytogenes strains from other noninfectious Listeria species DNA by using an inlA gene DNA probe [20]. Radhakrishnan et al. detected L. monocytogenes using monoclonal antibody deposited on an Au electrode with the help of electrochemical impedance spectroscopy. The results obtained showed that the sensitivity of the biosensor was 0.825 and 1.129 kΩ cm2/(CFU/mL) with 5 CFU/mL detection limit and 4 CFU/mL for working solutions and filtered tomato extract, respectively. Change in impedance due to non-specific binding was measured using a control antibody for glyceraldehyde 3-phosphate dehydrogenase (GADPH) and found to be insignificant [21]. Vanegas et al. developed a fast, reproducible, and sensitive electrochemical biosensor for determination of Listeria spp. depending upon the specific binding of InlA aptamers to internalins in the cell membrane of the target bacteria. Conjugated nanomaterial platforms based on reduced graphene oxide and nanoplatinum were deposited onto Pt/Ir electrodes that enhance the electrochemical transduction during the recognition events. These results exhibited that the aptasensor detected Listeria concentrations as low as 100 CFU/mL in <3 h (including incubation time and data analysis). Hence, this aptasensor has provided a new way for quick monitoring of Listeria monocytogenes in food products [22]. Nordin et al. detected Vibrio parahaemolyticus in diseased samples electrochemically using a screen-printed carbon electrode. The polylactide-stabilized gold nanoparticles were used

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for functionalization of the electrode, whereas methylene blue acted as a redox indicator. The biosensor has the ability to recognize precise complement, noncomplement, and mismatched sequences. The calibration range of the biosensor was analyzed in the range of 2.0  108–2.0  1013 M and 2.16 pM limit of detection. Detection of presence of Vibrio parahaemolyticus in spiked and unspiked samples of fresh cockles illustrated its simplicity and transportability, and hence, the device act as a significant screening device [23]. Pandey et al. have developed a marker free electrochemical biosensor for the efficient determination of diseases causing microorganisms. The biosensor was constructed by interaction of graphene with interdigitated microelectrodes of capacitors that were bio functionalized with E. coli O157:H7 targeted antibodies for detection of E. coli. Changes in the capacitance of capacitors showed that graphene based capacitors were specific for E. coli O157:H7 strain. The sensitivity of the biosensor was between 10 and 100 cells/mL. Hence, the biosensor provides multiple advantages like rapid, sensitive, specific, and in situ bacterial detection, free from use of chemical intermediaries, and is a useful strategy for determination of diverse nature of pathogens [24]. Ahari et al. constructed a potentiometric nanobiosensor technique by immobilizing the specific prototype of exotoxins of Staphylococcus aureus exotoxin. The biosensor was analyzed in terms of its impact of optimum pH on the S. aureus exotoxin nanobiosensor, which can be achieved by diluting the solutions of NaOH and HNO3, which were used for determining the higher and lower pH levels, correspondingly. This potentiometric nanobiosensor was able to sense an exotoxin density up to 10–3 M at 68 nm of synthesized molecularly imprinted polymer (MIP) during the initial 32 days of the experimentation (from a total of 56 days). Difference in the potential was stable at pH range of 5.0–8.5 and ideal temperature range of 15–25°C. Thus, pH and temperature play an important role in the precision of a potentiometric nanobiosensor for determining exotoxin of S. aureus [25].

31.5.3 Electrochemical Immunosensor Immunosensors are a component of the biosensor technology field with a diverse, wide, and multidisciplinary nature. They are basically designed to identify the direct binding of an antibody to an analyte to form an immunocomplex at the electrode surface. The basic principle behind the electrochemical immunosensor is to detect changes in the surface potential and oxidation state of an electro-active species. Electrochemical immunosensors provide extremely precise, rapid, and economical determination of pathogens with the potential to transform analytical procedures within the food industry. Detection of specific molecular recognition of an antigen by its antibodies is highly important in clinical diagnostics, and has the potential to provide quick and very sensitive HCV core antigen detection due to sensitivity, quick response, and inexpensive nanostructured metal oxides, which are the preferable choices to fabricate the sensor [26] (Scheme 31.2).

31.5.3.1 Recent application progress in different type of foods: l

Ding et al. developed an electrochemical immunosensor for the detection of severe pathogen, that is, Toxoplasma gondii (Tg). Herein this approach antibody against the pathogen was immobilized onto quartz crystal balance. The sensor was analyzed by the formation of precipitate which was formed upon interaction of pathogen with the antibody. In this

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+I +I +I +I

y od tib ation An 2. obiliz ified d m im n mo rode o lect e

Abs/NMs/bare electrode 1. Drop deposition of nanomaterial 3.

Bare electrode

NMs/bare electrode

Current (I)

Ab coupled with NMs by ionic and hydrophobic interactions

Ag a

dd

Potential (E/V)

ed

4. Electrochemical immunosensor for the detection of Ag-Ab

Ag-Ab interactions

Scheme 31.2 Schematic representation of electrochemical immunosensor.

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method, change in resistance was detected. The developed sensor has wide linear range and low detection limit [27]. Huang et al. have constructed a new impedimetric-based biosensor by immobilizing antibody against the pathogen. The immobilization matrix used was functionalized iron oxide nanoparticles as nanoparticles mediates the fast electron transfer which leads to the sensitive and rapid detection of Campylobacter jejuni. The change in resistance was proportional to the increased concentration of pathogen. The sensor showed wide linear range and low detection limit [28]. Tang et al. have developed a immune-based sensor for the detection of Staphylococcal enterotoxin B in sample of food. Herein this approach the immobilized matrix was Horseradish peroxidase tagged nanoslica-doped MWCNT which amplified the sensing signal. The developed sensor showed a wide linear range and have very low detection limit. The developed sensor was employed in spiked real samples. Samples used were apple juice, soymilk, and pork food. The developed sensor was in good relation with the conventional method [6]. Wan et al. have constructed a sensor by employing antibody against the sulfur-reducing bacteria. Electrodeposition technique was employed to develop sensing interface. The sensor was evaluated by electrochemical impedance technique and cyclic voltammetery. A wide linear range with good r value was reported in the present technique. The sensor was also found to be more specific toward the pathogen as sensor showed almost negligible response toward other pathogens [29]. Kanayeva et al. have synthesized magnetic nanoparticles and were used as sensing matrix. These nanoparticles were also functionalized for covalent coupling with the antibody. For covalent coupling, nanoparticles and antibody are modified with the streptavidin and biotin molecules. As both molecules showed high attraction toward each other. The sensor was

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found to be highly specific toward the Listerai as it showed almost insignificant response toward other pathogens [30]. Afonso et al. have constructed antigen-antibody-based biosensor for the detection of Salmonella sp. by employing gold nanoparticles. The immunosensor was found to have very low detection limit and wide linear range. The biosensor was also employed for the determination of Salmonella in spiked skimmed milk. The recovery was also found to be very satisfactory in the developed biosensor [31]. Kozitsina et al. have constructed an immunoassay platform for the specific detection of E. coli. Herein this approach, antibodies are immobilized onto the surface of electrode and the attached antibodies were able to detect the bacteria tagged magnetic nanoparticles. The developed immunosensor offered many advantageous features such as specificity, sensitivity, wide linear range, low detection limit, and good recovery values. The sensor was also employed for the determination of bacteria in real as well as spiked mixtures [32]. Masdor et al. developed a immunosensor based on direct and sandwich immunoassay for the specific detection of Campylobacter sp. The pathogen is mainly responsible for food poisoning. Herein this approaches both polyclonal and monoclonal antibodies were employed. In this method gold nanoparticles were employed for the detection of specific bacteria. The LOD of the developed sensor was found to be very low. The sensor was found to be highly sensitive and specific toward the detection of Campylobacter sp. This method did not require pre-enrichment method which leads to reduce in response time [33]. Wang et al. have constructed an electrochemical immunosensor by immobilizing a conjugate of gold nanoparticles (AuNPs) and polyclonal antibodies which formed a complex to produce MNP-targeted-AuNP sandwich. Analyzing the amount of AuNPs by the electrochemical method, the existence of target bacteria was determined. Results obtained have revealed a sensitivity of 101 colony forming units per milliliter (CFU/mL), 101–106 CFU/ mL linear range, 101 CFU/mL LOD, calibration range 101–106 CFU/mL having response time 45 min [34]. Tam et al. have investigated the Vibrio cholerae O1 using cerium oxide nanowire-based immunosensor. In this approach, electrochemical impedance spectroscopy was used for the detection of V. cholerae. This immunosensor was found to be highly efficient and sensitive [35]. Talan et al. developed an FTO-AuNPs-based electrochemical nanosensor to detect chlorpyrifos. In the working environment, the FTO-based nanosensor exhibited better sensitivity, and the chlorpyrifos concentration detecting range was found in between 1 fM and 1 μM with 10 fM limit of detection (LOD) up to 10 fM. Hence, the limit of detection of FTO-AuNPs biosensor for chlorpyrifos was found to be in standard and in real samples up to 10 nM for apple and cabbage, 50 nM for pomegranate [36]. Altintas et al. have developed a microfluidic-based electrochemical biosensor for the detection of E. coli. The presence of E. coli sensed by standard and nanomaterial amplified immu noassays was found in the concentration ranges of 0.99  104–3.98  109 CFU/mL and 10–3.97  107 CFU/mL with detection limits of 1.99  104 CFU/mL and 50 CFU/mL, respectively. Cross-reactivity studies have also been performed by monitoring Shigella, Salmonella spp., S. typhimurium and S. aureus, on E. coli specific antibody surface that renders the exceptional specificity of immunoassays [37].

31.5.4 Electrochemical Aptasensor Aptasensors are a novel biosensor in which aptamers are acting as bio-recognition elements. These sensors generate signal in the form of physical, chemical, electrical, or

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Paper electrode

1. Surface Working region modified with of bare paper- nanostructures based electrode

CV studies

NS/bare electrode

3. Electrochemical aptasensor for the detection of food pathogen

2. Aptamers immobilization onto modified electrode

Aptamer (probe)/NS/bare electrode

s. aureus (target)/ probe/NS/bare

Scheme 31.3 Schematic representation of electrochemical aptasensor.

optical change to determine the quantity of analyte in the samples when a suitable analyte interacts with an aptamer confined at the sensor substrate [38]. Electrochemical aptasensors have been developed by the immobilization of aptamers and recognition elements onto an electrode through electrochemical activity (Scheme 31.3). Nanomaterials have aroused interest as they enhance the sensitivity and stability of the biosensors due to their exceptional optical and electrical properties. Initially, under optimum conditions, the aptamers are confined at the electrode surface, which results in alteration in configuration of immobilized aptamers due to reaction of aptamers with the target analyte. Hence, changes in electrochemical signal in terms of potential, current, conductance, or impedance are recorded. This change in analytical signal could be used for sensing the target analyte. Such aptasensors provide high sensitivity, inexpensive, and unique specificity of aptamers with the target analyte [39]. Traditional potentiometric biosensors for bacteria detection require labeling of biorecognition elements. However, the interaction of pathogens with the bio-recognition component does not produce an electrochemical signal and hence it is necessary to use markers for the detection of changed potential. Zelada-Guillen and co-workers have constructed a marker independent potentiometric sensor for S. typhi detection through

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charge transfer ability of single-walled carbon nanotubes. Single-walled carbon nanotubes are used in aptasensor construction by immobilizing the functionalized aptamer at the surface of carbon nanotubes via a self-assembled monolayer. As the S. typhi bounded with aptamers led to alteration in the conformation of immobilized aptamers due to which phosphate group detached from the surface of carbon nanotubes, consequently there was a change in potential of the electrode. The biosensor exhibited a linear concentration range of 0.2–103 CFU/mL. This method has also been employed in detection of E. coli CECT 675 in milk and apple juice with a limit of detection 6 and 26 CFU/mL, respectively.

31.5.4.1 Recent application progress in different type of foods: l

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Giamberardino et al., using SELEX (Systematic Evolution of Ligands by EXponential enrichment), have fabricated DNA aptamers against murine norovirus (MNV). AG3 is the constituent of nine folds of SELEX, a promising aptamer exhibiting excellent specificity against MNV and for in vitro prepared capsids of human norovirus (HuNoV) outbreak strain (GII.3). The aptasensor has detected MNV with a limit of detection of about 180 virus entities and various probable site-directed applications. Thus, aptamers and the aptasensors have analyzed the fast detection and recognition of noroviruses in ecological as well as in medical samples [40]. Abbaspour et al. have detected S. aureus by designing an electrochemical dual aptasensor sandwich assay. The dynamic range of electrochemical immunosensor was 10 to 1  106 CFU/mL and 1.0 CFU/mL (S/N ¼ 3) detection limit. This aptasensor was also used for analyzing the interfering bacteria. The immunosensor was used for detecting the S. aureus in the real samples. The results were correlated with the experimental studies by plate counting method and thus the biosensor showed better consistency [41]. Jia et al. designed an aptasensor for the determination of S. aureus by using electrochemical impedance spectroscopy (EIS). The aptasensor was fabricated by conjugating singlestranded DNA with a nanocomposite fabricated from reduced graphene oxide (rGO) and gold nanoparticles (AuNPs). The aptasensor was applied for the detection of bacteria in the calibration range of 10–106 CFU/mL and resulted in a detection limit of 10 CFU/mL (S/N ¼ 3). The comparative standard divergence of S. aureus detection was equal to 4.3% (105 CFU/mL, n ¼ 7). Beside, its sensitivity, the aptamers-based sensor has showed unique selectivity as compared to other pathogens [42]. Ma et al. designed an electrochemical biosensor for the detection of Salmonella by using a Salmonella-specific recognition aptamer. As more Salmonella were introduced into the reaction system, the current produced between the electrode and electrolyte declined and resulted in detection limit <3 CFU mL. This novel method was specific, rapid, and applied for detection of Salmonella in a real sample [43]. Wang et al. constructed a novel electrochemical aptasensor for fast and sensitive determination of E. coli O157:H7 using immune magnetic nanoparticles (MNPs) assisted the precise separation of targeted bacteria; urease enzyme was used for urea for augmentation of the impedance signals and the PCB gold electrode for analyzing changed impedance. This proposed aptasensor based on MNPs was able to detect E. coli as low as 101 CFU/mL in 3 h, and the average recovery of E. coli in the spiked pasteurized milk was 99%. Thus, the aptasensor has been employed for detecting potential foodborne pathogens as the sensor has exhibited minimum response time, unique sensitivity, and minimum cost [44].

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Chand and Neethirajan developed an electrochemical aptasensor for norovirus sensing. This microfluidic aptasensor was analyzed by differential pulse voltammetry with limit of detection 100 pM and linear concentration detecting range from 100 pM to 3.5 nM. The proposed microfluidic aptasensor was practically used for the point-of-care one-step detection of norovirus in clinical samples [45].

31.5.5 Various foodborne pathogens and safety concerns There is diversity of microorganisms like fungi, bacteria, and viruses that spoil our food and are responsible for inducing several foodborne diseases and, consequently, food poisoning [46]. Microbes that cause foodborne diseases are known as foodborne pathogens and are a serious health threat in both developing and developed countries. Hence, food safety issues have attracted the attention of the worldwide scientific community for rapid-changing food recipes and food habits in the present as they have caused several fatal diseases. Foodborne ailments are due to pathogens, toxins, and other contaminants that represent a serious threat to human health [47]. Significant funds have been spent on analysis and on preventive measures, which is one of the considerable causes of food industry failures [48]. Table 31.1 shows different pathogens, their sources, and symptoms observed upon infection. Table 31.1 A tabular representation of various pathogens their sources and symptoms observed upon infection

Bacteria

Pathogens

Source

Symptoms

ØSalmonella nontyphoidal

Raw meats, raw milk, undercooked eggs, poultry, spices, contaminated water, vegetables, fruits Vegetable salads, nonveg items, diary products Vegetables, nonveg items, milk and its products such as butter, cheese, etc. Undercooked and raw vegetables or nonveg items, unpasteurized milk and milk products Sauces, processed foods such as soups, pickles Undercooked and raw vegetables or nonveg

Cramps, loose motion, severe fever, nausea

ØShigella spp. ØListeria monocytogenes

ØShiga-toxinproducing Escherichia coli (STEC) O157:H7 ØBacillus cereus ØCampylobacter spp.

Cramps, bloody loose motion, severe fever, nausea Feebleness, severe fever, neck stiffness, nausea Bloody diarrhea, cramps, stomachache, nausea Diarrhea, cramps, stomachache, nausea Bloody diarrhea, cramps, stomachache, nausea Continued

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Table 31.1 Continued Pathogens

ØClostridium perfringens ØStaphylococcus aureus ØYersinia enterocolitica

Parasitic

Viruses

Source items, unpasteurized milk and milk products Poultry and most of nonveg items Vegetable and nonveg salads, milk products such as butter, pastries, cake, and sandwich Seafood, poultry, unpasteurized milk, meat (pork, beef, lamb, etc.)

ØGiardia lamblia

Undercooked pork, lamb, or wild game

ØCryptosporidium parvum

Unprocessed foods, raw milk, meat and shellfish and fresh fruits and vegetables

ØNorovirus (Norwalk virus)

Ready to cook or eat foods, shell fish, food items contaminated with the vomit or feces Fishes from polluted water and food infected with feces or vomit, undercooked or raw food

ØHepatitis A virus

Symptoms

Cramps and diarrhea Diarrhea, cramps, stomachache, nausea, loss of appetite, and fever Enterocolitis, acute diarrhea, mesenteric lymphadenitis, pseudoappendicitis and terminal ileitis Fatigue, diarrhea or greasy stools, vomiting, weight loss, bloating and abdominal pain, excessive gas, headaches, abdominal pain and loss of appetite Watery diarrhea, nausea, vomiting, dehydration, loss of appetite, stomach cramps or pain and fever Severe diarrhea, abdominal pain, nausea Diarrhea, stomach ache, pain in head, nausea, loss in appetite

31.5.5.1 Salmonella l

Ma et al. have constructed a biosensor using aptamer as biological recognition element. They found that aptasensor has many advantageous features as compared to other sensors. In this approach, when there was increased concentration of Salmonella then the current response as found to be decreased. The sensor was also employed for the detection of Salmonella in real samples [43].

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Jia et al. have employed the reduced graphene oxide/carboxylated multiwalled carbon nanotubes onto the surface of electrode. Herein this approach, aptamer was employed as biological recognition element. Aptamer has the potential to directly recognize the pathogen. In this amino modified aptamer was immobilized onto the surface of electrode so that if forms covalent coupling with the –COOH group of multiwalled carbon nanotube. The biosensor showed very less detection limit and wide linear range [42]. Dastider et al. designed a microfluidic-based biosensor which is highly sensitive and specific for the detection of Salmonella sp. Authors have discussed the advantages approach of microfluidic-based device over nonfluidic devices. The developed biosensor showed very fast result and there is no requirement of any preenrichment. The detection limit of the microfluidic-based sensor found to be very less as compared to the nonmicrofluidic-based biosensor. The sensitivity and impedance performance of the microfluidic-based biosensor also get improved [49]. Stefano et al. reviewed the construction of various electrochemical biosensors such as genosensors, immune-sensors, etc. All biosensors are related with the fats and sensitive detection of Salmonella in food samples [50]. Li
ebana et al. highlightened the construction of various types of biosensors such as nucleic acid-based sensors, antibody-based sensors, and phagosensors. Various fast and sensitive methods were developed in order to detect the presence of Salmonella sp. Various new innovative technologies are developed in terms of biological recognition elements, transducers, incorporation of microfluidic approach, economical platforms, point of need devices, and multiplexed detection of various pathogens at one platform [51]. Liu et al. have designed a new biosensor for rapid detection of Salmonella sp. in samples of food by employing multiwalled carbon nanotube/chitosan/peroxidase decorated screen printed electrode and antibody against salmonella is immobilized on CN membrane. The limit of detection of sensor was found to be very low due to the use of employed matrix and response obtained was very fast. The designed portable device is found to be effective in the detection of Salmonella sp. in food samples [52]. An immunological sandwich ELISA test was designed employing antibody against Salmonella tagged with peroxidase enzyme. Tetramethylbenzidine dihydrochloride was employed as mediator. The developed system was sensitive for the detection of Salmonella sp. [53].

31.5.5.2 Listeria l

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Wu et al. constructed an electrochemical sensor for the detection of LLO toxin genes in food products or organisms by immobilizing single-stranded oligonucleotides onto gold electrodes by means of mercaptan of the DNA bases with N-hydroxysulfosuccinimide (NHS) and N-(3-dimethylamion)propyl-N0 -ethyl carbodiimide hydrochloride (EDC) for activation [17]. Bifulco et al. designed an electrochemical genosensor by immobilizing single-stranded DNA for the detection of the inlA gene in Listeria strains without labeling the target DNA. The immobilized single-stranded DNA was hybridized to its complementary DNA sequences to form double-stranded DNA on the gold surface. As a result, significant decline in current of the voltammogram (DPV) was recorded [20]. Radhakrishnan et al. developed an impedometric sensor by immobilizing mouse monoclonal antibody onto Au electrode for the detection of L. monocytogenes. The sensor yielded sensitivities of 0.825 and 1.129 kΩ cm2/(CFU/mL) and detection limits 5 and 4 CFU/mL for ideal solutions and filtered tomato extract, respectively. The impedance change from

808

l

l

Evaluation Technologies for Food Quality

nonspecific binding was quantified with a control antibody for glyceraldehyde 3-phosphate dehydrogenase (GADPH), and found to be insignificant [21]. Vanegas et al. detected Listeria spp. based on the selective binding of InlA aptamers to internalins in the cell membrane of the target bacteria by developing an electrochemical biosensor. This electrochemical biosensor was rapid, accurate, and sensitive. Herein conjugated nanomaterials of reduced graphene oxide and nanoplatinum were confined at the surface of Pt/Ir electrodes and, consequently, thereby increasing the electrochemical signals. The sensor showed a limit of detection of 100 CFU/mL in <3 h [22]. Chen et al. have fabricated a unique optical biosensor for the fast and efficient determination of Listeria monocytogenes. This proposed biosensor exhibited a detection limit of 1.0  102 CFU/mL with an average recovery of 95.1% for Listeria in the spiked lettuce samples. This biosensor has exhibited a number of advantages such as facile, economic, sensitive, and very low limit of detection during analysis of foodborne pathogen [54].

31.5.5.3 Staphylococcus aureus l

Bhardwaj et al. designed an ultrasensitive biosensor precisely for S. aureus. Bacteriophage was interfaced with a water-dispersible and environmentally stable metal-organic framework (MOF), NH2-MIL-5(Fe). The MOF-bacteriophage biosensor was used for sensitive detection of S. aureus in both artificial and real samples by photoluminescence quenching phenomena. The limit of detection of proposed biosensor was 31 CFU/mL and detecting concentration range 40 to 4  108 CFU/mL [55].

31.5.5.4 Norovirus l

l

Hong et al. developed a highly sensitive, discriminatory, and fast electrochemical biosensor using nanostructured gold electrode combined with concanavalin A (ConA) for the detection of NoV. Cyclic voltammetric experiments have shown a linear concentration range of NoV in between 102 and 106 copies/mL, reduced response time (1 h), and better detection limit (35 copies/mL). Furthermore, the biosensor has exhibited exceptional reproducibility (RSD ¼ 4.38%), as well as thermal stability (declines of 2.5% and 3.1% at 4°C and 25°C, respectively) [56]. Hwang et al. have fabricated marker-free electrochemical biosensor for the investigation of human infecting norovirus. This biosensor was highly sensitive as it uses specific peptide for scrutinizing the norovirus. The biosensor exhibited 99.8 nM limit of detection for recombinant noroviral capsid proteins (rP2) and 7.8 copies/mL for human norovirus [57].

31.5.6 Low cost platforms for portable NP-based detection Transducers used for the fabrication of cost-effective NP-based sensors are either screen-printed electrodes or paper. Screen-printed electrodes are low in cost and are employed in artistic applications and for design electronic circuits [58]. Biosensors based on screen printed technology have exhibited many advantages such as not being expansive, easy modification, ease of reproducibility, and sensitivity. Beside carbon nanomaterials, several other nanomaterials like CeO2, Au, Ag, and ZnO

Sensors for quality analysis of foods

809

NPs have been used in sensing components, by using functional electrodes that enlarge the surface area, produce unique catalytic activity, and augment the electrochemical signals. Screen-printed technology also includes the use of NPs or nanotubes in composition of screen-printed inks [59] and sometimes nanomaterials are drop-casted in DMF/ water or deposited at the surface of working electrodes electrochemically [60]. Nanomaterials used in the fabrication of SPCE electrodes have offered several advantages, which include [61]: (a) matrix for biomolecules immobilization and increased consistency and bioactivity; (b) acting as mediator for promoting electron transfer reactions, reduced optimum potential, and minimum interference problems, better sensitivity and selectivity; (c) marker for electrochemical striping techniques which produce an electrochemical signal; and (d) as a catalyst which augments the electrochemical signal. Alternatively, paper has created interest as a sensing platform, as it is a facile, cheap, and copious material. For instance, paper bioassays, such as specific paper prepared by photolithography for determination of glucose and bovine serum albumin [62], inkjet-printed paper fluidic immunochemical sensing device [63], aptamer NP-based lateral flow devices for analysis of DNA sequences [64], and inkjet-printed enzyme sensors have been used for the determination of bisphenol A in field samples [65, 66]. These paper-based devices can be connected with colorimetric [67] and electrochemical [68, 69] detecting procedures. The application of paper-based electrochemical sensors has been illustrated in detecting precise components in diverse fields like environment protection and medicine, as well as in food safety measurements [70]. Baxter and co-workers have designed a facile and inexpensive method by fabricating gold electrodes on paper via a camera flash sintering procedure [71]. Recently, a portable and reagentless NP-based paper-based sensor for the detection of oxidase enzyme substrates, such as glucose [72] and polyphenols, in food samples such as wild mushrooms, wine, juice, and green tea [73] using electrochemical and surface chelating properties of nanoceria. Determination of an analyte has also been carried out by measuring the intensity of color change of the particles after product formation during an enzymatic reaction (H2O2) of the polyphenol.

31.6

Use of electrosensors for chemical and biological contamination

Analyte Acetylcholinesterase (AChE) Chlorpyrifos E. coli

Detection principle Electrochemical Immunosensor Electrochemical immunosensor Electrochemical biochips

Detection limit 8.0 pM 10 fM 500 colony forming units (CFU)

Dynamic range 10 pM to 4 nM 1 fM to 1 μM 400, 80, 500 CFU

Refs. [53] [36] [74]

Continued

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Evaluation Technologies for Food Quality

Continued Detection principle

Analyte Pork meatball Hydrogen peroxide in fruit juice Methyl parathion

31.7

Electrochemical sensor Amperometric sensor Electrochemical sensor

Detection limit 6, 4, and 0.23 μg/ mL 0.99–28 μM, 28–510 μM, and 0.51–1.8 mM 5 nM–2780 μM

Dynamic range

Refs.

4 μg/mL

[75]

0.27 μM

[76]

0.5 nM

[77]

Conclusions and future direction

Analytical devices based NPs and nanostructures have significantly enhanced detection capabilities. Sensors based on these nanomaterials have exhibited unique sensitivity and low detection limits. Some detection procedures use sensitive reagents and multiple steps, which prolong the analysis time, increase the price, and make field implementation complicated. Transformation of this method for improving food quality requires validation via traditional techniques, investigation of real samples, and cautious assessment of interferences. Furthermore, issues have been raised regarding the toxicity of nanomaterials. Investigation of food is a very complicated problem because of the intrinsic complexity of these samples. Most biosensors require sample-preparation steps. Integration of sample extraction and separation units with the sensing devices would improve the portability of sensing devices in field use. Storage of food for prolonged time is another big issue for systems that incorporate biological recognition elements like enzymes and antibodies. Furthermore, it is necessary to attain high specificity, to reduce background signals, and to reduce related false positive results. Miniaturization, mechanization, ability to detect multiple analytes, and attempts to minimize the price per assay are a few recent issues in this area. Use of paper to fabricate sensors is cost-effective and has ease of portability; however, their performance for sensing complex food samples and requirement of sample pretreatment are yet to be demonstrated. In future, research should be focused on improving food quality by means of food packaging that will reduce the chances of contamination of processed and stored food. Improved portability of the sensor can be achieved by connectivity and combination with widely used communicating tools including cell-phones and tablets. Fabrication of cybersensors for scrutinizing food is in the development stage and opens lots of opportunity for future research.

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