Search results for: enzymatic biosensor
Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 509

Search results for: enzymatic biosensor

449 Immuno-field Effect Transistor Using Carbon Nanotubes Network – Based for Human Serum Albumin Highly Sensitive Detection

Authors: Muhamad Azuddin Hassan, Siti Shafura Karim, Ambri Mohamed, Iskandar Yahya

Abstract:

Human serum albumin plays a significant part in the physiological functions of the human body system (HSA).HSA level monitoring is critical for early detection of HSA-related illnesses. The goal of this study is to show that a field effect transistor (FET)-based immunosensor can assess HSA using high aspect ratio carbon nanotubes network (CNT) as a transducer. The CNT network were deposited using air brush technique, and the FET device was made using a shadow mask process. Field emission scanning electron microscopy and a current-voltage measurement system were used to examine the morphology and electrical properties of the CNT network, respectively. X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy were used to confirm the surface alteration of the CNT. The detection process is based on covalent binding interactions between an antibody and an HSA target, which resulted in a change in the manufactured biosensor's drain current (Id).In a linear range between 1 ng/ml and 10zg/ml, the biosensor has a high sensitivity of 0.826 mA (g/ml)-1 and a LOD value of 1.9zg/ml.HSA was also identified in a genuine serum despite interference from other biomolecules, demonstrating the CNT-FET immunosensor's ability to quantify HSA in a complex biological environment.

Keywords: carbon nanotubes network, biosensor, human serum albumin

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448 Low-Complex, High-Fidelity Two-Grades Cyclo-Olefin Copolymer (COC) Based Thermal Bonding Technique for Sealing a Thermoplastic Microfluidic Biosensor

Authors: Jorge Prada, Christina Cordes, Carsten Harms, Walter Lang

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The development of microfluidic-based biosensors over the last years has shown an increasing employ of thermoplastic polymers as constitutive material. Their low-cost production, high replication fidelity, biocompatibility and optical-mechanical properties are sought after for the implementation of disposable albeit functional lab-on-chip solutions. Among the range of thermoplastic materials on use, the Cyclo-Olefin Copolymer (COC) stands out due to its optical transparency, which makes it a frequent choice as manufacturing material for fluorescence-based biosensors. Moreover, several processing techniques to complete a closed COC microfluidic biosensor have been discussed in the literature. The reported techniques differ however in their implementation, and therefore potentially add more or less complexity when using it in a mass production process. This work introduces and reports results on the application of a purely thermal bonding process between COC substrates, which were produced by the hot-embossing process, and COC foils containing screen-printed circuits. The proposed procedure takes advantage of the transition temperature difference between two COC grades foils to accomplish the sealing of the microfluidic channels. Patterned heat injection to the COC foil through the COC substrate is applied, resulting in consistent channel geometry uniformity. Measurements on bond strength and bursting pressure are shown, suggesting that this purely thermal bonding process potentially renders a technique which can be easily adapted into the thermoplastic microfluidic chip production workflow, while enables a low-cost as well as high-quality COC biosensor manufacturing process.

Keywords: biosensor, cyclo-olefin copolymer, hot embossing, thermal bonding, thermoplastics

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447 A Theoretical Modelling and Simulation of a Surface Plasmon Resonance Biosensor for the Detection of Glucose Concentration in Blood and Urine

Authors: Natasha Mandal, Rakesh Singh Moirangthem

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The present work reports a theoretical model to develop a plasmonic biosensor for the detection of glucose concentrations in human blood and urine as the abnormality of glucose label is the major cause of diabetes which becomes a life-threatening disease worldwide. This study is based on the surface plasmon resonance (SPR) sensor applications which is a well-established, highly sensitive, label-free, rapid optical sensing tool. Here we have introduced a sandwich assay of two dielectric spacer layers of MgF2 and BaTiO3which gives better performance compared to commonly used SiO2 and TiO2 dielectric spacers due to their low dielectric loss and higher refractive index. The sensitivity of our proposed sensor was found as 3242 nm/RIU approximately, with an excellent linear response of 0.958, which is higher than the conventional single-layer Au SPR sensor. Further, the sensitivity enhancement is also optimized by coating a few layers of two-dimensional (2D) nanomaterials (e.g., Graphene, h-BN, MXene, MoS2, WS2, etc.) on the sensor chip. Hence, our proposed SPR sensor has the potential for the detection of glucose concentration in blood and urine with enhanced sensitivity and high affinity and could be utilized as a reliable platform for the optical biosensing application in the field of medical diagnosis.

Keywords: biosensor, surface plasmon resonance, dielectric spacer, 2D nanomaterials

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446 Biosensor: An Approach towards Sustainable Environment

Authors: Purnima Dhall, Rita Kumar

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Introduction: River Yamuna, in the national capital territory (NCT), and also the primary source of drinking water for the city. Delhi discharges about 3,684 MLD of sewage through its 18 drains in to the Yamuna. Water quality monitoring is an important aspect of water management concerning to the pollution control. Public concern and legislation are now a day’s demanding better environmental control. Conventional method for estimating BOD5 has various drawbacks as they are expensive, time-consuming, and require the use of highly trained personnel. Stringent forthcoming regulations on the wastewater have necessitated the urge to develop analytical system, which contribute to greater process efficiency. Biosensors offer the possibility of real time analysis. Methodology: In the present study, a novel rapid method for the determination of biochemical oxygen demand (BOD) has been developed. Using the developed method, the BOD of a sample can be determined within 2 hours as compared to 3-5 days with the standard BOD3-5day assay. Moreover, the test is based on specified consortia instead of undefined seeding material therefore it minimizes the variability among the results. The device is coupled to software which automatically calculates the dilution required, so, the prior dilution of the sample is not required before BOD estimation. The developed BOD-Biosensor makes use of immobilized microorganisms to sense the biochemical oxygen demand of industrial wastewaters having low–moderate–high biodegradability. The method is quick, robust, online and less time consuming. Findings: The results of extensive testing of the developed biosensor on drains demonstrate that the BOD values obtained by the device correlated with conventional BOD values the observed R2 value was 0.995. The reproducibility of the measurements with the BOD biosensor was within a percentage deviation of ±10%. Advantages of developed BOD biosensor • Determines the water pollution quickly in 2 hours of time; • Determines the water pollution of all types of waste water; • Has prolonged shelf life of more than 400 days; • Enhanced repeatability and reproducibility values; • Elimination of COD estimation. Distinctiveness of Technology: • Bio-component: can determine BOD load of all types of waste water; • Immobilization: increased shelf life > 400 days, extended stability and viability; • Software: Reduces manual errors, reduction in estimation time. Conclusion: BiosensorBOD can be used to measure the BOD value of the real wastewater samples. The BOD biosensor showed good reproducibility in the results. This technology is useful in deciding treatment strategies well ahead and so facilitating discharge of properly treated water to common water bodies. The developed technology has been transferred to M/s Forbes Marshall Pvt Ltd, Pune.

Keywords: biosensor, biochemical oxygen demand, immobilized, monitoring, Yamuna

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445 Multi-Walled Carbon Nanotubes Doped Poly (3,4 Ethylenedioxythiophene) Composites Based Electrochemical Nano-Biosensor for Organophosphate Detection

Authors: Navpreet Kaur, Himkusha Thakur, Nirmal Prabhakar

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One of the most publicized and controversial issue in crop production is the use of agrichemicals- also known as pesticides. This is evident in many reports that Organophosphate (OP) insecticides, among the broad range of pesticides are mainly involved in acute and chronic poisoning cases. Therefore, detection of OPs is very necessary for health protection, food and environmental safety. In our study, a nanocomposite of poly (3,4 ethylenedioxythiophene) (PEDOT) and multi-walled carbon nanotubes (MWCNTs) has been deposited electrochemically onto the surface of fluorine doped tin oxide sheets (FTO) for the analysis of malathion OP. The -COOH functionalization of MWCNTs has been done for the covalent binding with amino groups of AChE enzyme. The use of PEDOT-MWCNT films exhibited an excellent conductivity, enables fast transfer kinetics and provided a favourable biocompatible microenvironment for AChE, for the significant malathion OP detection. The prepared PEDOT-MWCNT/FTO and AChE/PEDOT-MWCNT/FTO nano-biosensors were characterized by Fourier transform infrared spectrometry (FTIR), Field emission-scanning electron microscopy (FE-SEM) and electrochemical studies. Electrochemical studies were done using Cyclic Voltammetry (CV) or Differential Pulse Voltammetry (DPV) and Electrochemical Impedance Spectroscopy (EIS). Various optimization studies were done for different parameters including pH (7.5), AChE concentration (50 mU), substrate concentration (0.3 mM) and inhibition time (10 min). The detection limit for malathion OP was calculated to be 1 fM within the linear range 1 fM to 1 µM. The activity of inhibited AChE enzyme was restored to 98% of its original value by 2-pyridine aldoxime methiodide (2-PAM) (5 mM) treatment for 11 min. The oxime 2-PAM is able to remove malathion from the active site of AChE by means of trans-esterification reaction. The storage stability and reusability of the prepared nano-biosensor is observed to be 30 days and seven times, respectively. The application of the developed nano-biosensor has also been evaluated for spiked lettuce sample. Recoveries of malathion from the spiked lettuce sample ranged between 96-98%. The low detection limit obtained by the developed nano-biosensor made them reliable, sensitive and a low cost process.

Keywords: PEDOT-MWCNT, malathion, organophosphates, acetylcholinesterase, nano-biosensor, oxime (2-PAM)

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444 Highly Specific DNA-Aptamer-Based Electrochemical Biosensor for Mercury (II) and Lead (II) Ions Detection in Water Samples

Authors: H. Abu-Ali, A. Nabok, T. Smith

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Aptamers are single-strand of DNA or RNA nucleotides sequence which is designed in vitro using selection process known as SELEX (systematic evolution of ligands by exponential enrichment) were developed for the selective detection of many toxic materials. In this work, we have developed an electrochemical biosensor for highly selective and sensitive detection of Hg2+ and Pb2+ using a specific aptamer probe (SAP) labelled with ferrocene (or methylene blue) in (5′) end and the thiol group at its (3′) termini, respectively. The SAP has a specific coil structure that matching with G-G for Pb2+ and T-T for Hg2+ interaction binding nucleotides ions, respectively. Aptamers were immobilized onto surface of screen-printed gold electrodes via SH groups; then the cyclic voltammograms were recorded in binding buffer with the addition of the above metal salts in different concentrations. The resulted values of anode current increase upon binding heavy metal ions to aptamers and analyte due to the presence of electrochemically active probe, i.e. ferrocene or methylene blue group. The correlation between the anodic current values and the concentrations of Hg2+ and Pb2+ ions has been established in this work. To the best of our knowledge, this is the first example of using a specific DNA aptamers for electrochemical detection of heavy metals. Each increase in concentration of 0.1 μM results in an increase in the anode current value by simple DC electrochemical test i.e (Cyclic Voltammetry), thus providing an easy way of determining Hg2+ and Pb2+concentration.

Keywords: aptamer, based, biosensor, DNA, electrochemical, highly, specific

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443 Biosensor Design through Molecular Dynamics Simulation

Authors: Wenjun Zhang, Yunqing Du, Steven W. Cranford, Ming L. Wang

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The beginning of 21st century has witnessed new advancements in the design and use of new materials for biosensing applications, from nano to macro, protein to tissue. Traditional analytical methods lack a complete toolset to describe the complexities introduced by living systems, pathological relations, discrete hierarchical materials, cross-phase interactions, and structure-property dependencies. Materiomics – via systematic molecular dynamics (MD) simulation – can provide structure-process-property relations by using a materials science approach linking mechanisms across scales and enables oriented biosensor design. With this approach, DNA biosensors can be utilized to detect disease biomarkers present in individuals’ breath such as acetone for diabetes. Our wireless sensor array based on single-stranded DNA (ssDNA)-decorated single-walled carbon nanotubes (SWNT) has successfully detected trace amount of various chemicals in vapor differentiated by pattern recognition. Here, we present how MD simulation can revolutionize the way of design and screening of DNA aptamers for targeting biomarkers related to oral diseases and oral health monitoring. It demonstrates great potential to be utilized to build a library of DNDA sequences for reliable detection of several biomarkers of one specific disease, and as well provides a new methodology of creating, designing, and applying of biosensors.

Keywords: biosensor, DNA, biomarker, molecular dynamics simulation

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442 A Stepwise Approach for Piezoresistive Microcantilever Biosensor Optimization

Authors: Amal E. Ahmed, Levent Trabzon

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Due to the low concentration of the analytes in biological samples, the use of Biological Microelectromechanical System (Bio-MEMS) biosensors for biomolecules detection results in a minuscule output signal that is not good enough for practical applications. In response to this, a need has arisen for an optimized biosensor capable of giving high output signal in response the detection of few analytes in the sample; the ultimate goal is being able to convert the attachment of a single biomolecule into a measurable quantity. For this purpose, MEMS microcantilevers based biosensors emerged as a promising sensing solution because it is simple, cheap, very sensitive and more importantly does not need analytes optical labeling (Label-free). Among the different microcantilever transducing techniques, piezoresistive based microcantilever biosensors became more prominent because it works well in liquid environments and has an integrated readout system. However, the design of piezoresistive microcantilevers is not a straightforward problem due to coupling between the design parameters, constraints, process conditions, and performance. It was found that the parameters that can be optimized to enhance the sensitivity of Piezoresistive microcantilever-based sensors are: cantilever dimensions, cantilever material, cantilever shape, piezoresistor material, piezoresistor doping level, piezoresistor dimensions, piezoresistor position, Stress Concentration Region's (SCR) shape and position. After a systematic analyzation of the effect of each design and process parameters on the sensitivity, a step-wise optimization approach was developed in which almost all these parameters were variated one at each step while fixing the others to get the maximum possible sensitivity at the end. At each step, the goal was to optimize the parameter in a way that it maximizes and concentrates the stress in the piezoresistor region for the same applied force thus get the higher sensitivity. Using this approach, an optimized sensor that has 73.5x times higher electrical sensitivity (ΔR⁄R) than the starting sensor was obtained. In addition to that, this piezoresistive microcantilever biosensor it is more sensitive than the other similar sensors previously reported in the open literature. The mechanical sensitivity of the final senior is -1.5×10-8 Ω/Ω ⁄pN; which means that for each 1pN (10-10 g) biomolecules attach to this biosensor; the piezoresistor resistivity will decrease by 1.5×10-8 Ω. Throughout this work COMSOL Multiphysics 5.0, a commercial Finite Element Analysis (FEA) tool, has been used to simulate the sensor performance.

Keywords: biosensor, microcantilever, piezoresistive, stress concentration region (SCR)

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441 Production of Camel Nanobodies against of Anti-Morphine-3-Glucuronide for the Development of a Biosensor for Detecting Illicit Drug

Authors: Shirin Jalili, Sadegh Hasannia, Hadi Shirzad, Afshin Khara

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Morphine is one of the most medicinally important analgesics and narcotics. Structurally, it is classified as an alkaloid because of the presence of nitrogen. Its structure is similar to that of codeine, thebaine, and heroin. An immunoassay to accurately discriminate between these analogous alkaloids would be highly beneficial. A key factor for such an assay is specificity with high sensitivity, which is totally dependent on the antibody employed. However, most antibodies against haptens are polyclonal serum antibodies that exhibit significant cross-reactivities with closely related compounds. The camel-derived single-chain antibody fragments (VHH) are the smallest molecules with antigen-binding capacity, possessing unique properties compared to other conventional antibodies. In this study, a library containing the VHH genes of a camel immunized with with morphine conjugated BSA following phage display technology was generated. By screening the camel-derived variable region of the heavy chain cDNA phage display library with the ability to bind the desired hapten, we obtained some nanobodies that recognize this hapten. Phage display expression of the Nbs from this library and pannings against this hapten resulted in a clear enrichment of four distinct Nb-displaying phages with specificity for morphine that could be a potential target site for the development of new strategies for the development of a biosensor for detecting illicit drug.

Keywords: phage display, nanobody, Morphine-3, glucuronide, ELISA, biosensor

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440 Impedimetric Phage-Based Sensor for the Rapid Detection of Staphylococcus aureus from Nasal Swab

Authors: Z. Yousefniayejahr, S. Bolognini, A. Bonini, C. Campobasso, N. Poma, F. Vivaldi, M. Di Luca, A. Tavanti, F. Di Francesco

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Pathogenic bacteria represent a threat to healthcare systems and the food industry, because their rapid detection remains challenging. Electrochemical biosensors are gaining prominence as a novel technology for the detection of pathogens due to intrinsic features such as low cost, rapid response time, and portability, which make them a valuable alternative to traditional methodologies. These sensors use biorecognition elements that are crucial for the identification of specific bacteria. In this context, bacteriophages are promising tools for their inherent high selectivity towards bacterial hosts, which is of fundamental importance when detecting bacterial pathogens in complex biological samples. In this study, we present the development of a low-cost and portable sensor based on the Zeno phage for the rapid detection of Staphylococcus aureus. Screen-printed gold electrodes functionalized with the Zeno phage were used, and electrochemical impedance spectroscopy was applied to evaluate the change of the charge transfer resistance (Rct) as a result of the interaction with S. aureus MRSA ATCC 43300. The phage-based biosensor showed a linear range from 101 to 104 CFU/mL with a 20-minute response time and a limit of detection (LOD) of 6 CFU/mL under physiological conditions. The biosensor’s ability to recognize various strains of staphylococci was also successfully demonstrated in the presence of clinical isolates collected from different geographic areas. Assays using S. epidermidis were also carried out to verify the species-specificity of the phage sensor. We only observed a remarkable change of the Rct in the presence of the target S. aureus bacteria, while no substantial binding to S. epidermidis occurred. This confirmed that the Zeno phage sensor only targets S. aureus species within the genus Staphylococcus. In addition, the biosensor's specificity with respect to other bacterial species, including gram-positive bacteria like Enterococcus faecium and the gram-negative bacterium Pseudomonas aeruginosa was evaluated, and a non-significant impedimetric signal was observed. Notably, the biosensor successfully identified S. aureus bacterial cells in a complex matrix such as a nasal swab, opening the possibility of its use in a real-case scenario. We diluted different concentrations of S. aureus from 108 to 100 CFU/mL with a ratio of 1:10 in the nasal swap matrices collected from healthy donors. Three different sensors were applied to measure various concentrations of bacteria. Our sensor indicated high selectivity to detect S. aureus in biological matrices compared to time-consuming traditional methods, such as enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR), and radioimmunoassay (RIA), etc. With the aim to study the possibility of using this biosensor to address the challenge associated with pathogen detection, ongoing research is focused on the assessment of the biosensor’s analytical performances in different biological samples and the discovery of new phage bioreceptors.

Keywords: electrochemical impedance spectroscopy, bacteriophage, biosensor, Staphylococcus aureus

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439 Development of Biosensor Chip for Detection of Specific Antibodies to HSV-1

Authors: Zatovska T. V., Nesterova N. V., Baranova G. V., Zagorodnya S. D.

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In recent years, biosensor technologies based on the phenomenon of surface plasmon resonance (SPR) are becoming increasingly used in biology and medicine. Their application facilitates exploration in real time progress of binding of biomolecules and identification of agents that specifically interact with biologically active substances immobilized on the biosensor surface (biochips). Special attention is paid to the use of Biosensor analysis in determining the antibody-antigen interaction in the diagnostics of diseases caused by viruses and bacteria. According to WHO, the diseases that are caused by the herpes simplex virus (HSV), take second place (15.8%) after influenza as a cause of death from viral infections. Current diagnostics of HSV infection include PCR and ELISA assays. The latter allows determination the degree of immune response to viral infection and respective stages of its progress. In this regard, the searches for new and available diagnostic methods are very important. This work was aimed to develop Biosensor chip for detection of specific antibodies to HSV-1 in the human blood serum. The proteins of HSV1 (strain US) were used as antigens. The viral particles were accumulated in cell culture MDBK and purified by differential centrifugation in cesium chloride density gradient. Analysis of the HSV1 proteins was performed by polyacrylamide gel electrophoresis and ELISA. The protein concentration was measured using De Novix DS-11 spectrophotometer. The device for detection of antigen-antibody interactions was an optoelectronic two-channel spectrometer ‘Plasmon-6’, using the SPR phenomenon in the Krechman optical configuration. It was developed at the Lashkarev Institute of Semiconductor Physics of NASU. The used carrier was a glass plate covered with 45 nm gold film. Screening of human blood serums was performed using the test system ‘HSV-1 IgG ELISA’ (GenWay, USA). Development of Biosensor chip included optimization of conditions of viral antigen sorption and analysis steps. For immobilization of viral proteins 0.2% solution of Dextran 17, 200 (Sigma, USA) was used. Sorption of antigen took place at 4-8°C within 18-24 hours. After washing of chip, three times with citrate buffer (pH 5,0) 1% solution of BSA was applied to block the sites not occupied by viral antigen. It was found direct dependence between the amount of immobilized HSV1 antigen and SPR response. Using obtained biochips, panels of 25 positive and 10 negative for the content of antibodies to HSV-1 human sera were analyzed. The average value of SPR response was 185 a.s. for negative sera and from 312 to. 1264 a.s. for positive sera. It was shown that SPR data were agreed with ELISA results in 96% of samples proving the great potential of SPR in such researches. It was investigated the possibility of biochip regeneration and it was shown that application of 10 mM NaOH solution leads to rupture of intermolecular bonds. This allows reuse the chip several times. Thus, in this study biosensor chip for detection of specific antibodies to HSV1 was successfully developed expanding a range of diagnostic methods for this pathogen.

Keywords: biochip, herpes virus, SPR

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438 Development of Technologies for Biotransformation of Aquatic Biological Resources for the Production of Functional, Specialized, Therapeutic, Preventive, and Microbiological Products

Authors: Kira Rysakova, Vitaly Novikov

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An improved method of obtaining enzymatic collagen hydrolysate from the tissues of marine hydrobionts is proposed, which allows to obtain hydrolysate without pre-isolation of pure collagen. The method can be used to isolate enzymatic collagen hydrolysate from the waste of industrial processing of Red King crab and non-traditional objects - marine holothurias. Comparative analysis of collagen hydrolysates has shown the possibility of their use in a number of nutrient media, but this requires additional optimization of their composition and biological tests on wide sets of test strains of microorganisms.

Keywords: collagen hydrolysate, marine hydrobionts, red king crab, marine holothurias, enzymes, exclusive HPLC

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437 Bienzymatic Nanocomposites Biosensors Complexed with Gold Nanoparticles, Polyaniline, Recombinant MN Peroxidase from Corn, and Glucose Oxidase to Measure Glucose

Authors: Anahita Izadyar

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Using a recombinant enzyme derived from corn and a simple modification, we are fabricating a facile, fast, and cost-beneficial novel biosensor to measure glucose. We are applying Plant Produced Mn Peroxidase (PPMP), glucose oxidase (GOx), polyaniline (PANI) as conductive polymer and gold nanoparticles (AuNPs) on Au electrode using electrochemical response to detect glucose. We applied the entrapment method of enzyme composition, which is generally used to immobilize conductive polymer and facilitate electron transfer from the enzyme oxidation-reduction center to the sample solution. In this work, the oxidation of glucose on the modified gold electrode was quantified with Linear Sweep Voltammetry(LSV). We expect that the modified biosensor has the potential for monitoring various biofluids.

Keywords: plant-produced manganese peroxidase, enzyme-based biosensors, glucose, modified gold nanoparticles electrode, polyaniline

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436 Enzymatic Repair Prior To DNA Barcoding, Aspirations, and Restraints

Authors: Maxime Merheb, Rachel Matar

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Retrieving ancient DNA sequences which in return permit the entire genome sequencing from fossils have extraordinarily improved in recent years, thanks to sequencing technology and other methodological advances. In any case, the quest to search for ancient DNA is still obstructed by the damage inflicted on DNA which accumulates after the death of a living organism. We can characterize this damage into three main categories: (i) Physical abnormalities such as strand breaks which lead to the presence of short DNA fragments. (ii) Modified bases (mainly cytosine deamination) which cause errors in the sequence due to an incorporation of a false nucleotide during DNA amplification. (iii) DNA modifications referred to as blocking lesions, will halt the PCR extension which in return will also affect the amplification and sequencing process. We can clearly see that the issues arising from breakage and coding errors were significantly decreased in recent years. Fast sequencing of short DNA fragments was empowered by platforms for high-throughput sequencing, most of the coding errors were uncovered to be the consequences of cytosine deamination which can be easily removed from the DNA using enzymatic treatment. The methodology to repair DNA sequences is still in development, it can be basically explained by the process of reintroducing cytosine rather than uracil. This technique is thus restricted to amplified DNA molecules. To eliminate any type of damage (particularly those that block PCR) is a process still pending the complete repair methodologies; DNA detection right after extraction is highly needed. Before using any resources into extensive, unreasonable and uncertain repair techniques, it is vital to distinguish between two possible hypotheses; (i) DNA is none existent to be amplified to begin with therefore completely un-repairable, (ii) the DNA is refractory to PCR and it is worth to be repaired and amplified. Hence, it is extremely important to develop a non-enzymatic technique to detect the most degraded DNA.

Keywords: ancient DNA, DNA barcodong, enzymatic repair, PCR

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435 Physicochemical Properties and Thermal Inactivation of Polyphenol Oxidase of African Bush Mango (Irvingia Gabonensis) Fruit

Authors: Catherine Joke Adeseko

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Enzymatic browning is an economically important disorder that degrades organoleptic properties and prevent the consumer from purchasing fresh fruit and vegetables. Prevention and control of enzymatic browning in fruit and its product is imperative. Therefore, this study sought to investigate the catalytic effect of polyphenol oxidase (PPO) in the adverse browning of African bush mango (Irvingia gabonensis) fruit peel and pulp. PPO was isolated and purified, and its physicochemical properties, such as the effect of pH with SDS, temperature, and thermodynamic studies, which invariably led to thermal inactivation of purified PPO at 80 °C, were evaluated. The pH and temperature optima of PPO were found at 7.0 and 50, respectively. There was a gradual increase in the activity of PPO as the pH increases. However, the enzyme exhibited a higher activity at neutral pH 7.0, while enzymatic inhibition was observed at acidic region, pH 2.0. The presence of SDS at pH 5.0 downward was found to inhibit the activity of PPO from the peel and pulp of I. gabonensis. The average value of enthalpy (ΔH), entropy (ΔS), and Gibbs free energy (ΔG) obtained at 20 min of incubation and temperature 30 – 80 °C were respectively 39.93 kJ.mol-1, 431.57 J.mol-1 .K-1 and -107.99 kJ.mol-1 for peel PPO, and 37.92 kJ.mol-1, -442.51J.mol-1.K-1, and -107.22 kJ.mol-1 for pulp PPO. Thermal inactivation of PPO from I. gabonensis exhibited a reduction in catalytic activity as the temperature and duration of heat inactivation increases using catechol, reflected by an increment in k value. The half-life of PPO (t1/2) decreases as the incubation temperature increases due to the instability of the enzyme at high temperatures and was higher in pulp than peel. Both D and Z values decrease with increase in temperature. The information from this study suggests processing parameters for controlling PPO in the potential industrial application of I. gabonensis fruit in order to prolong the shelf-life of this fruit for maximum utilization.

Keywords: enzymatic, browning, characterization, activity

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434 The Methods of Immobilization of Laccase for Direct Transfer in an Enzymatic Fuel Cell

Authors: Afshin Farahbakhsh, Hoda Khodadadi

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In this paper, we compare five methods of biological fuel cell fabrication by combining a Shewanella oneidensis microbial anode and a laccase-modified air-breathing cathode. As a result of biofuel cell laccase with graphite nanofibers, carbon surface (PAMAN) on the pt/hpg electrode, graphite sheets MWCNT and with (PG) and (MWCNT) showed, respectively. Describes methods for creating controllable and reproducible bio-anodes and demonstrates the versatility of hybrid biological fuel cells. The laccase-based biocathodes prepared either with the crude extract or with the purified enzyme can provide electrochemically active and stable biomaterials. The laccase-based biocathodes prepared either with the crude extract or with the purified enzyme can provide electrochemically active and stable biomaterials. When the device was fed with transdermal extracts, containing only 30μM of glucose, the average peak power was proportionally lower (0.004mW). The result of biofuel cell with graphite nanofibers showed the enzymatic fuel cell reaches 0.5 V at open circuit voltage with both, ethanol and methanol and the maximum current density observed for E2electrode was 228.94mAcm.

Keywords: enzymatic electrode, fuel cell, immobilization, laccase

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433 Non Enzymatic Electrochemical Sensing of Glucose Using Manganese Doped Nickel Oxide Nanoparticles Decorated Carbon Nanotubes

Authors: Anju Joshi, C. N. Tharamani

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Diabetes is one of the leading cause of death at present and remains an important concern as the prevalence of the disease is increasing at an alarming rate. Therefore, it is crucial to diagnose the accurate levels of glucose for developing an efficient therapeutic for diabetes. Due to the availability of convenient and compact self-testing, continuous monitoring of glucose is feasible nowadays. Enzyme based electrochemical sensing of glucose is quite popular because of its high selectivity but suffers from drawbacks like complicated purification and immobilization procedures, denaturation, high cost, and low sensitivity due to indirect electron transfer. Hence, designing a robust enzyme free platform using transition metal oxides remains crucial for the efficient and sensitive determination of glucose. In the present work, manganese doped nickel oxide nanoparticles (Mn-NiO) has been synthesized onto the surface of multiwalled carbon nanotubes using a simple microwave assisted approach for non-enzymatic electrochemical sensing of glucose. The morphology and structure of the synthesized nanostructures were characterized using scanning electron microscopy (SEM) and X-Ray diffraction (XRD). We demonstrate that the synthesized nanostructures show enormous potential for electrocatalytic oxidation of glucose with high sensitivity and selectivity. Cyclic voltammetry and square wave voltammetry studies suggest superior sensitivity and selectivity of Mn-NiO decorated carbon nanotubes towards the non-enzymatic determination of glucose. A linear response between the peak current and the concentration of glucose has been found to be in the concentration range of 0.01 μM- 10000 μM which suggests the potential efficacy of Mn-NiO decorated carbon nanotubes for sensitive determination of glucose.

Keywords: diabetes, glucose, Mn-NiO decorated carbon nanotubes, non-enzymatic

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432 Two-Step Patterning of Microfluidic Structures in Paper by Laser Cutting and Wax Printing for Mass Fabrication of Biosensor

Authors: Bong Keun Kang, Sung Suk Oh, Jeong-Woo Sohn, Jong-Ryul Choi, Young Ho Kim

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In this paper, we describe two-step micro-pattering by using laser cutting and wax printing. Wax printing is performed only on the bridges for hydrophobic barriers. We prepared 405nm blue-violet laser module and wax pencil module. And, this two modules combine x-y plot. The hollow microstructure formed by laser patterning define the hydrophilic flowing paths. However, bridges are essential to avoid the cutting area being the island. Through the support bridges, microfluidic solution spread out to the unnecessary areas. Chromatography blotting paper was purchased from Whatman. We used 20x20 cm and 46x57 cm of chromatography blotting paper. Axis moving speed of x-y plot was the main parameter of optimization. For aligning between the two patterning, the paper sheet was taped at the bottom. After the two-step patterning, temperature curing step was done at 110-130 °C. The resolution of the fabrication and the potential of the multiplex detection were investigated.

Keywords: µPADs, microfluidic, biosensor, mass-fabrication

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431 Candida antarctica Lipase-B Catalyzed Alkaline-Hydrolysis of Some Aryl-Alkyl Acetate in Non-Aqueous Media

Authors: M. Merabet-Khelassi, Z. Houiene, L. Aribi-Zouioueche, O. Riant

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Lipases (EC.3.1.1.3) are efficient biotools widely used for their remarkable chemo-, regio- and enantio-selectivity, especially, in kinetic resolution of racemates. They offer access to a large panel of enantiopure building blocks, such as secondary benzylic alcohols, commonly used as synthetic intermediates in pharmaceutical and agrochemical industries. Due to the stability of lipases in both water and organic solvents poor in water, they are able to catalyze both transesterifications of arylalkylcarbinols and hydrolysis of their corresponding acetates. The use of enzymatic hydrolysis in aqueous media still limited. In this presentation, we expose a practical methodology for the preparation of optically enriched acetates using a Candida antarctica lipase B-catalyzed hydrolysis in non-aqueous media in the presence of alkaline carbonate salts. The influence of several parameters which can intervene on the enzymatic efficiency such as the impact of the introduction of the carbonates salts, its amount and the nature of the alkaline earth metal are discussed. The obtained results show that the use of sodium carbonate with CAL-B enhances drastically both reactivity and selectivity of this immobilized lipase. In all cases, the resulting alcohols and remaining acetates are obtained in high ee values (up to > 99 %), and the selectivities reach (E > 500).

Keywords: alkaline-hydrolysis, enzymatic kinetic resolution, lipases, arylalkylcarbinol, non-aqueous media

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430 Reagentless Detection of Urea Based on ZnO-CuO Composite Thin Film

Authors: Neha Batra Bali, Monika Tomar, Vinay Gupta

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A reagentless biosensor for detection of urea based on ZnO-CuO composite thin film is presented in following work. Biosensors have immense potential for varied applications ranging from environmental to clinical testing, health care, and cell analysis. Immense growth in the field of biosensors is due to the huge requirement in today’s world to develop techniques which are both cost effective and accurate for prevention of disease manifestation. The human body comprises of numerous biomolecules which in their optimum levels are essential for functioning. However mismanaged levels of these biomolecules result in major health issues. Urea is one of the key biomolecules of interest. Its estimation is of paramount significance not only for healthcare sector but also from environmental perspectives. If level of urea in human blood/serum is abnormal, i.e., above or below physiological range (15-40mg/dl)), it may lead to diseases like renal failure, hepatic failure, nephritic syndrome, cachexia, urinary tract obstruction, dehydration, shock, burns and gastrointestinal, etc. Various metal nanoparticles, conducting polymer, metal oxide thin films, etc. have been exploited to act as matrix to immobilize urease to fabricate urea biosensor. Amongst them, Zinc Oxide (ZnO), a semiconductor metal oxide with a wide band gap is of immense interest as an efficient matrix in biosensors by virtue of its natural abundance, biocompatibility, good electron communication feature and high isoelectric point (9.5). In spite of being such an attractive candidate, ZnO does not possess a redox couple of its own which necessitates the use of electroactive mediators for electron transfer between the enzyme and the electrode, thereby causing hindrance in realization of integrated and implantable biosensor. In the present work, an effort has been made to fabricate a matrix based on ZnO-CuO composite prepared by pulsed laser deposition (PLD) technique in order to incorporate redox properties in ZnO matrix and to utilize the same for reagentless biosensing applications. The prepared bioelectrode Urs/(ZnO-CuO)/ITO/glass exhibits high sensitivity (70µAmM⁻¹cm⁻²) for detection of urea (5-200 mg/dl) with high stability (shelf life ˃ 10 weeks) and good selectivity (interference ˂ 4%). The enhanced sensing response obtained for composite matrix is attributed to the efficient electron exchange between ZnO-CuO matrix and immobilized enzymes, and subsequently fast transfer of generated electrons to the electrode via matrix. The response is encouraging for fabricating reagentless urea biosensor based on ZnO-CuO matrix.

Keywords: biosensor, reagentless, urea, ZnO-CuO composite

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429 Biosensor Technologies in Neurotransmitters Detection

Authors: Joanna Cabaj, Sylwia Baluta, Karol Malecha

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Catecholamines are vital neurotransmitters that mediate a variety of central nervous system functions, such as motor control, cognition, emotion, memory processing, and endocrine modulation. Dysfunctions in catecholamine neurotransmission are induced in some neurologic and neuropsychiatric diseases. Changeable neurotransmitters level in biological fluids can be a marker of several neurological disorders. Because of its significance in analytical techniques and diagnostics, sensitive and selective detection of neurotransmitters is increasingly attracting a lot of attention in different areas of bio-analysis or biomedical research. Recently, optical techniques for the detection of catecholamines have attracted interests due to their reasonable cost, convenient control, as well as maneuverability in biological environments. Nevertheless, with the observed need for a sensitive and selective catecholamines sensor, the development of a convenient method for this neurotransmitter is still at its basic level. The manipulation of nanostructured materials in conjunction with biological molecules has led to the development of a new class of hybrid-modified enzymatic sensors in which both enhancement of charge transport and biological activity preservation may be obtained. Immobilization of biomaterials on electrode surfaces is the crucial step in fabricating electrochemical as well as optical biosensors and bioelectronic devices. Continuing systematic investigation in manufacturing of enzyme–conducting sensitive systems, here is presented a convenient fluorescence as well as electrochemical sensing strategy for catecholamines detection.

Keywords: biosensors, catecholamines, fluorescence, enzymes

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428 Effect of Enzymatic Hydrolysis and Ultrasounds Pretreatments on Biogas Production from Corn Cob

Authors: N. Pérez-Rodríguez, D. García-Bernet, A. Torrado-Agrasar, J. M. Cruz, A. B. Moldes, J. M. Domínguez

Abstract:

World economy is based on non-renewable, fossil fuels such as petroleum and natural gas, which entails its rapid depletion and environmental problems. In EU countries, the objective is that at least 20% of the total energy supplies in 2020 should be derived from renewable resources. Biogas, a product of anaerobic degradation of organic substrates, represents an attractive green alternative for meeting partial energy needs. Nowadays, trend to circular economy model involves efficiently use of residues by its transformation from waste to a new resource. In this sense, characteristics of agricultural residues (that are available in plenty, renewable, as well as eco-friendly) propitiate their valorisation as substrates for biogas production. Corn cob is a by-product obtained from maize processing representing 18 % of total maize mass. Corn cob importance lies in the high production of this cereal (more than 1 x 109 tons in 2014). Due to its lignocellulosic nature, corn cob contains three main polymers: cellulose, hemicellulose and lignin. Crystalline, highly ordered structures of cellulose and lignin hinders microbial attack and subsequent biogas production. For the optimal lignocellulose utilization and to enhance gas production in anaerobic digestion, materials are usually submitted to different pretreatment technologies. In the present work, enzymatic hydrolysis, ultrasounds and combination of both technologies were assayed as pretreatments of corn cob for biogas production. Enzymatic hydrolysis pretreatment was started by adding 0.044 U of Ultraflo® L feruloyl esterase per gram of dry corncob. Hydrolyses were carried out in 50 mM sodium-phosphate buffer pH 6.0 with a solid:liquid proportion of 1:10 (w/v), at 150 rpm, 40 ºC and darkness for 3 hours. Ultrasounds pretreatment was performed subjecting corn cob, in 50 mM sodium-phosphate buffer pH 6.0 with a solid: liquid proportion of 1:10 (w/v), at a power of 750W for 1 minute. In order to observe the effect of the combination of both pretreatments, some samples were initially sonicated and then they were enzymatically hydrolysed. In terms of methane production, anaerobic digestion of the corn cob pretreated by enzymatic hydrolysis was positive achieving 290 L CH4 kg MV-1 (compared with 267 L CH4 kg MV-1 obtained with untreated corn cob). Although the use of ultrasound as the only pretreatment resulted detrimentally (since gas production decreased to 244 L CH4 kg MV-1 after 44 days of anaerobic digestion), its combination with enzymatic hydrolysis was beneficial, reaching the highest value (300.9 L CH4 kg MV-1). Consequently, the combination of both pretreatments improved biogas production from corn cob.

Keywords: biogas, corn cob, enzymatic hydrolysis, ultrasound

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427 Optimization of Enzymatic Hydrolysis of Cooked Porcine Blood to Obtain Hydrolysates with Potential Biological Activities

Authors: Miguel Pereira, Lígia Pimentel, Manuela Pintado

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Animal blood is a major by-product of slaughterhouses and still represents a cost and environmental problem in some countries. To be eliminated, blood should be stabilised by cooking and afterwards the slaughterhouses must have to pay for its incineration. In order to reduce the elimination costs and valorise the high protein content the aim of this study was the optimization of hydrolysis conditions, in terms of enzyme ratio and time, in order to obtain hydrolysates with biological activity. Two enzymes were tested in this assay: pepsin and proteases from Cynara cardunculus (cardosins). The latter has the advantage to be largely used in the Portuguese Dairy Industry and has a low price. The screening assays were carried out in a range of time between 0 and 10 h and using a ratio of enzyme/reaction volume between 0 and 5%. The assays were performed at the optimal conditions of pH and temperature for each enzyme: 55 °C at pH 5.2 for cardosins and 37 °C at pH 2.0 for pepsin. After reaction, the hydrolysates were evaluated by FPLC (Fast Protein Liquid Chromatography) and tested for their antioxidant activity by ABTS method. FPLC chromatograms showed different profiles when comparing the enzymatic reactions with the control (no enzyme added). The chromatogram exhibited new peaks with lower MW that were not present in control samples, demonstrating the hydrolysis by both enzymes. Regarding to the antioxidant activity, the best results for both enzymes were obtained using a ratio enzyme/reactional volume of 5% during 5 h of hydrolysis. However, the extension of reaction did not affect significantly the antioxidant activity. This has an industrial relevant aspect in what concerns to the process cost. In conclusion, the enzymatic blood hydrolysis can be a better alternative to the current elimination process allowing to the industry the reuse of an ingredient with biological properties and economic value.

Keywords: antioxidant activity, blood, by-products, enzymatic hydrolysis

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426 Comparative Electrochemical Studies of Enzyme-Based and Enzyme-less Graphene Oxide-Based Nanocomposite as Glucose Biosensor

Authors: Chetna Tyagi. G. B. V. S. Lakshmi, Ambuj Tripathi, D. K. Avasthi

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Graphene oxide provides a good host matrix for preparing nanocomposites due to the different functional groups attached to its edges and planes. Being biocompatible, it is used in therapeutic applications. As enzyme-based biosensor requires complicated enzyme purification procedure, high fabrication cost and special storage conditions, we need enzyme-less biosensors for use even in a harsh environment like high temperature, varying pH, etc. In this work, we have prepared both enzyme-based and enzyme-less graphene oxide-based biosensors for glucose detection using glucose-oxidase as enzyme and gold nanoparticles, respectively. These samples were characterized using X-ray diffraction, UV-visible spectroscopy, scanning electron microscopy, and transmission electron microscopy to confirm the successful synthesis of the working electrodes. Electrochemical measurements were performed for both the working electrodes using a 3-electrode electrochemical cell. Cyclic voltammetry curves showed the homogeneous transfer of electron on the electrodes in the scan range between -0.2V to 0.6V. The sensing measurements were performed using differential pulse voltammetry for the glucose concentration varying from 0.01 mM to 20 mM, and sensing was improved towards glucose in the presence of gold nanoparticles. Gold nanoparticles in graphene oxide nanocomposite played an important role in sensing glucose in the absence of enzyme, glucose oxidase, as evident from these measurements. The selectivity was tested by measuring the current response of the working electrode towards glucose in the presence of the other common interfering agents like cholesterol, ascorbic acid, citric acid, and urea. The enzyme-less working electrode also showed storage stability for up to 15 weeks, making it a suitable glucose biosensor.

Keywords: electrochemical, enzyme-less, glucose, gold nanoparticles, graphene oxide, nanocomposite

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425 The Response of Adaptive Mechanism of Fluorescent Proteins from Coral Species and Target Cell Properties on Signalling Capacity as Biosensor

Authors: Elif Tugce Aksun Tumerkan

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Fluorescent proteins (FPs) have become very popular since green fluorescent protein discovered from crystal jellyfish. It is known that Anthozoa species have a wide range of chromophore organisms, and the initial crystal structure for non-fluorescent chromophores obtained from the reef-building coral has been determined. There are also differently coloured pigments in non-bioluminescent Anthozoa zooxanthellate and azooxanthellate which are frequently members of the GFP-like protein family. The development of fluorescent proteins (FPs) and their applications is an outstanding example of basic science leading to practical biotechnological and medical applications. Fluorescent proteins have several applications in science and are used as important indicators in molecular biology and cell-based research. With rising interest in cell biology, FPs have used as biosensor indicators and probes in pharmacology and cell biology. Using fluorescent proteins in genetically encoded metabolite sensors has many advantages than chemical probes for metabolites such as easily introduced into any cell or organism in any sub-cellular localization and giving chance to fixing to fluoresce of different colours or characteristics. There are different factors effects to signalling mechanism when they used as a biosensor. While there are wide ranges of research have been done on the significance and applications of fluorescent proteins, the cell signalling response of FPs and target cell are less well understood. In this study, it was aimed to clarify the response of adaptive mechanisms of coral species such as pH, temperature and symbiotic relationship and target cells properties on the signalling capacity. Corals are a rich natural source of fluorescent proteins that change with environmental conditions such as light, heat stress and injury. Adaptation mechanism of coral species to these types of environmental variations is important factor due to FPs properties have affected by this mechanism. Since fluorescent proteins obtained from nature, their own ecological property like the symbiotic relationship is observed very commonly in coral species and living conditions have the impact on FPs efficiency. Target cell properties also have an effect on signalling and visualization. The dynamicity of detector that used for reading fluorescence and the level of background fluorescence are key parameters for the quality of the fluorescent signal. Among the factors, it can be concluded that coral species adaptive characteristics have the strongest effect on FPs signalling capacity.

Keywords: biosensor, cell biology, environmental conditions, fluorescent protein, sea anemone

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424 Jamun Juice Extraction Using Commercial Enzymes and Optimization of the Treatment with the Help of Physicochemical, Nutritional and Sensory Properties

Authors: Payel Ghosh, Rama Chandra Pradhan, Sabyasachi Mishra

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Jamun (Syzygium cuminii L.) is one of the important indigenous minor fruit with high medicinal value. The jamun cultivation is unorganized and there is huge loss of this fruit every year. The perishable nature of the fruit makes its postharvest management further difficult. Due to the strong cell wall structure of pectin-protein bonds and hard seeds, extraction of juice becomes difficult. Enzymatic treatment has been commercially used for improvement of juice quality with high yield. The objective of the study was to optimize the best treatment method for juice extraction. Enzymes (Pectinase and Tannase) from different stains had been used and for each enzyme, best result obtained by using response surface methodology. Optimization had been done on the basis of physicochemical property, nutritional property, sensory quality and cost estimation. According to quality aspect, cost analysis and sensory evaluation, the optimizing enzymatic treatment was obtained by Pectinase from Aspergillus aculeatus strain. The optimum condition for the treatment was 44 oC with 80 minute with a concentration of 0.05% (w/w). At these conditions, 75% of yield with turbidity of 32.21NTU, clarity of 74.39%T, polyphenol content of 115.31 mg GAE/g, protein content of 102.43 mg/g have been obtained with a significant difference in overall acceptability.

Keywords: enzymatic treatment, Jamun, optimization, physicochemical property, sensory analysis

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423 Water Re-Use Optimization in a Sugar Platform Biorefinery Using Municipal Solid Waste

Authors: Leo Paul Vaurs, Sonia Heaven, Charles Banks

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Municipal solid waste (MSW) is a virtually unlimited source of lignocellulosic material in the form of a waste paper/cardboard mixture which can be converted into fermentable sugars via cellulolytic enzyme hydrolysis in a biorefinery. The extraction of the lignocellulosic fraction and its preparation, however, are energy and water demanding processes. The waste water generated is a rich organic liquor with a high Chemical Oxygen Demand that can be partially cleaned while generating biogas in an Upflow Anaerobic Sludge Blanket bioreactor and be further re-used in the process. In this work, an experiment was designed to determine the critical contaminant concentrations in water affecting either anaerobic digestion or enzymatic hydrolysis by simulating multiple water re-circulations. It was found that re-using more than 16.5 times the same water could decrease the hydrolysis yield by up to 65 % and led to a complete granules desegregation. Due to the complexity of the water stream, the contaminant(s) responsible for the performance decrease could not be identified but it was suspected to be caused by sodium, potassium, lipid accumulation for the anaerobic digestion (AD) process and heavy metal build-up for enzymatic hydrolysis. The experimental data were incorporated into a Water Pinch technology based model that was used to optimize the water re-utilization in the modelled system to reduce fresh water requirement and wastewater generation while ensuring all processes performed at optimal level. Multiple scenarios were modelled in which sub-process requirements were evaluated in term of importance, operational costs and impact on the CAPEX. The best compromise between water usage, AD and enzymatic hydrolysis yield was determined for each assumed contaminant degradations by anaerobic granules. Results from the model will be used to build the first MSW based biorefinery in the USA.

Keywords: anaerobic digestion, enzymatic hydrolysis, municipal solid waste, water optimization

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422 Biological Regulation of Endogenous Enzymatic Activity of Rainbow Trout (Oncorhynchus Mykiss) with Protease Inhibitors Chickpea in Model Systems

Authors: Delgado-Meza M., Minor-Pérez H.

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Protease is the generic name of enzymes that hydrolyze proteins. These are classified in the subgroup EC3.4.11-99X of the classification enzymes. In food technology the proteolysis is used to modify functional and nutritional properties of food, and in some cases this proteolysis may cause food spoilage. In general, seafood and rainbow trout have accelerated decomposition process once it has done its capture, due to various factors such as the endogenous enzymatic activity that can result in loss of structure, shape and firmness, besides the release of amino acid precursors of biogenic amines. Some studies suggest the use of protease inhibitors from legume as biological regulators of proteolytic activity. The enzyme inhibitors are any substance that reduces the rate of a reaction catalyzed by an enzyme. The objective of this study was to evaluate the reduction of the proteolytic activity of enzymes in extracts of rainbow trout with protease inhibitors obtained from chickpea flour. Different proportions of rainbow trout enzyme extract (75%, 50% and 25%) and extract chickpea enzyme inhibitors were evaluated. Chickpea inhibitors were obtained by mixing 5 g of flour in 30 mL of pH 7.0 phosphate buffer. The sample was centrifuged at 8000 rpm for 10 min. The supernatant was stored at -15°C. Likewise, 20 g of rainbow trout were ground in 20 mL of phosphate buffer solution at pH 7.0 and the mixture was centrifuged at 5000 rpm for 20 min. The supernatant was used for the study. In each treatment was determined the specific enzymatic activity with the technique of Kunitz, using hemoglobin as substrate for the enzymes acid fraction and casein for basic enzymes. Also biuret protein was quantified for each treatment. The results showed for fraction of basic enzymes in the treatments evaluated, that were inhibition of endogenous enzymatic activity. Inhibition values compared to control were 51.05%, 56.59% and 59.29% when the proportions of endogenous enzymes extract rainbow trout were 75%, 50% and 25% and the remaining volume used was extract with inhibitors. Treatments with acid enzymes showed no reduction in enzyme activity. In conclusion chickpea flour reduced the endogenous enzymatic activity of rainbow trout, which may favor its application to increase the half-life of this food. The authors acknowledge the funding provided by the CONACYT for the project 131998.

Keywords: rainbouw trout, enzyme inhibitors, proteolysis, enzyme activity

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421 In Vitro Digestibility of Grains and Straw of Seventeen Ecotypes of Bitter Vetch (Vicia ervilia) in the North of Morocco

Authors: Boukrouh Soumaya, Cabaraux Jean-François, Avril Claire, Noutfia Ali, Chentouf Mouad

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The introduction of marginal leguminous forage species in the diet of ruminants are of great importance. Bitter vetch is a good source of proteins, highly resistant against drought and poor soil conditions. Accordingly; two years field trials (2018/2019 and 2019-2020) were conducted to determine the digestibility of straw and grains of 17 promising bitter vetch ecotypes(Vicia ervilia) in the north of Morocco. In vitro dry and organic matter digestibility, gas production, and kinetics of fermentation of grains and straw were evaluated using gas production technique, pepsin-cellulase enzymatic digestibility of DM (CDDM)and OM (CDOM), as well as protease enzymatic CP degradation (CPD) and in vitro true digestibility, were performed using DAISYII Incubator. In vitro digestibility was performed using gas production method of (Menke et al., 1979) improved by Menke and Steingass (1988). Samples were incubated in glass syringes that contained rumen fluid and incubation solution that conserved in water bath in 39°C during 72 hours. Gas production was recorded after 2, 4, 8, 12, 24, 48, and 72 hours. Studied digestibility parameters were dry and organic matter digestibility, microbial biomass production, partitioning factor, and volatile fatty acids. Enzymatic dry matter digestibility was different (p < 0.05) among grains and straw for all ecotypes. It varied from 804.1 to 957.7 g/kg DM and 270.4 to 412.3 g/kg DM for grains and straw, respectively. Metabolizable energy varied between 11.7 to 14.3 MJ/kg DM and 2.6 to 5.0 MJ/kg DM for grains and straw, respectively. Potential gas production (A), the rate constants (c and d), and lag times of grains and straws from different bitter vetch ecotypes were different (p > 0.05). The results emphasized that in any evaluation of bitter vetch ecotypes, where straw of this legume seed is used as an animal feed, not only seed yield but also yield and quality of straw should be taken into consideration, particularly in areas where straw from this legume is considered as an important feedstuff for ruminants. Enzymatic digestibility was lower than in vitro digestibility by gaz production and by the DAISYII method because rumen fluid contains bacteria than increase digestibility. There was no difference between in vitro digestibility by gaz production and the DAISY II method. The DAISY II method can be used to increase labor efficiency in the in vitro DM digestibility analysis if gaz production is not necessary for analysis.

Keywords: bitter vetch, grains, straw, ecotype, in vitro digestibility, gaz production, enzymatic digestibility

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420 Meta-Review of Scholarly Publications on Biosensors: A Bibliometric Study

Authors: Nasrine Olson

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With over 70,000 scholarly publications on the topic of biosensors, an overview of the field has become a challenge. To facilitate, there are currently over 700 expert-reviews of publications on biosensors and related topics. This study focuses on these review papers in order to provide a Meta-Review of the area. This paper provides a statistical analysis and overview of biosensor-related review papers. Comprehensive searches are conducted in the Web of Science, and PubMed databases and the resulting empirical material are analyzed using bibliometric methods and tools. The study finds that the biosensor-related review papers can be categorized in five related subgroups, broadly denoted by (i) properties of materials and particles, (ii) analysis and indicators, (iii) diagnostics, (iv) pollutant and analytical devices, and (v) treatment/ application. For an easy and clear access to the findings visualization of clusters and networks of connections are presented. The study includes a temporal dimension and identifies the trends over the years with an emphasis on the most recent developments. This paper provides useful insights for those who wish to form a better understanding of the research trends in the area of biosensors.

Keywords: bibliometrics, biosensors, meta-review, statistical analysis, trends visualization

Procedia PDF Downloads 186