Immobilization Of Hemoglobin Research Articles

Electroconductive bioplatform based on dextrin for the immobilization of hemoglobin: Application for electrochemical monitoring of H2O2

A novel biodegradable dextrin-based nanocomposite, involving polypyrrole (PPy) and hydrophilic dextrin (Dex) (PPy@Dex) was prepared using in-situ radical chemical polymerization technique. The obtained PPy@Dex bionanocomposite was fully characterized by FT-IR, XRD, FESEM, and DSC methods. The exceptional properties such as biocompatibility, high surface area, the proper functional group on the surface, and outstanding electrical conductivity of synthesized bionanocomposite made it a superior candidate over biomolecules immobilization. Electrochemical observations revealed that the PPy@Dex-coated glassy carbon electrode (GCE) demonstrated improved performance, making it a suitable substrate for immobilizing hemoglobin (Hb) and constructing an efficient biosensor. The resulting biosensor, named Hb-PPy@Dex/GCE, exhibited high activity in the reduction of hydrogen peroxide (H2O2). Amperometric examinations demonstrated an extensive linear range from 2 to 350 μM for Hb-PPy@Dex/GCE. The detection limit of the proposed approach was calculated to be 0.54 μM, following the S/N = 3 protocol.

Immobilization of hemoglobin on MnCO3 sphere-loaded Au nanoparticles as highly efficient sensing platform towards hydrogen peroxide

In this paper, we report the synthesis of MnCO3−Au hybrid microspheres and their application on the electrochemical biosensing of hydrogen peroxide (H2O2) based on the immobilization of hemoglobin (Hb). The characterization of MnCO3−Au microspheres revealed that an abundance of Au nanoparticles (AuNPs) has been absorbed on the surface of the spherical MnCO3 by the electrostatic assembly. The combined unique properties of MnCO3−Au microspheres are beneficial for the realization of the direct electron transfer of Hb. Hb immobilized on the microspheres maintained its biological activity, showing a surface-controlled process with the heterogeneous electron transfer rate constant (k s) of 2.63 s−1. The fabricated biosensor displayed an excellent performance for the electrocatalytic reduction of H2O2. The linear range for the determination of H2O2 was from 0.06–40.0 μM with a detection limit of 0.015 µM (S/N = 3). The biosensor also exhibited high selectivity, good repeatability and long-term stability, which offers great potential for H2O2 detection in real sample analysis.

Withdrawal Notice: Immobilization of Hemoglobin onto Nanomesoporous MCM-41 and Its Catalytic Performance

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Bioelectrocatalysis and direct determination of H2O2 using the high-performance platform: chitosan nanofibers modified with SDS and hemoglobin

In the present work, a novel platform has been designed to fabricate an electrochemical sensor composed of chitosan nanofibers modified with sodium dodecyl sulfate, a negatively charged surfactant, and hemoglobin protein for direct and sensitive determination of hydrogen peroxide. Chitosan nanofibers were synthesized through the electrospinning process and characterized by Fourier transform infrared spectroscopy and scanning electron microscopy. The synthesized nanofibers provided a large surface for the adsorption of hemoglobin. Immobilization of hemoglobin on the fibers was investigated by cyclic voltammetry, and the modification steps were followed by electrochemical impedance spectroscopy. Under the optimal conditions, the fabricated biosensor was employed for electrocatalytic determination of hydrogen peroxide by differential pulse voltammetric and amperometric methods in the linear range 3.0–2940.0 µM (detection limit = 0.16 µM) and 1.0–201.0 µM (detection limit = 0.05 µM), respectively. The applicability of the constructed biosensor was also evaluated by hydrogen peroxide determination in real samples, and satisfactory recoveries were obtained which confirmed that the proposed method provides a simple and sensitive method for the hydrogen peroxide monitoring.

Development of Hydrogen Peroxide Biosensor Based on Immobilization of Hemoglobin on Screen-Printed Carbon Electrode Modified With Silver Nanoparticles

The direct electron transfer behavior of hemoglobin that is immobilized onto screen-printed carbon electrode (SPCE) modified with silver nanoparticles (AgNPs) and chitosan (CS) was studied in this work. Cyclic voltametry and spectrophotometry were used to characterize the hemoglobin (Hb) bioconjunction with AgNPs and CS. Results of the modified electrode showed quasi-reversible redox peaks with a formal potential of (-0.245V) versus Ag/AgCl in 0.1M phosphate buffer solution (PBS), pH7, at a scan rate of 0.1Vs-1. The charge transfer coefficient (α) was 0.48 and the apparent electron transfer rate constant (Ks) was 0.47s-1. The electrode was used as a hydrogen peroxide biosensor with a linear response over 3 to 240 µM and a detection limit of 0.6 µM. As a result, the modified biosensor here has exhibited a high sensitivity, good reproducibility and stability.

Simple and sensitive detection of acrylamide based on hemoglobin immobilization in carbon ionic liquid paste electrode

A simple and sensitive electrochemical biosensor for acrylamide (AA) detection based on hemoglobin (Hb) entrapped in ionic liquid-carbon paste was constructed. The Hb-carbon ionic liquid paste electrode (Hb/CILPE) was studied and characterized by scanning electron microscopy (SEM) and electrochemical method. The surface passivation of the Hb/CILPE could be achieved by the addition of acrylamide molecule, which caused the lower current response value owing to the adduct formed by acrylamide and Hb as a result of the reaction between the α-NH2 group and the N-terminal valine of Hb. The experimental conditions such as pH value of supporting electrolyte, pulse increment and concentrated time of AA was examined. Under the optimum conditions, the differential pulse voltammetry (DPV) responses of Hb/CILPE toward AA was evaluated in a good linear range from 1.0 × 10−11 to 1.0 × 10−5 M, with a detection limit of 5.0 × 10−12 M at S/N = 3. This biosensor can be successfully applied to the determination of AA in a real sample without complex pretreatment. Compared with other electrochemical methods for detecting AA, the prepared electrochemical biosensor exhibited advantages of simple preparation, easy surface renewal, better reproducibility and satisfactory stability.

Study on the immobilization of hemoglobin by nano mesoporous MCFs and its optical properties

Nano mesoporous molecular sieve MCFs (mesocellular foams) was synthesized successfully through hydrothermal method. Then bovine hemoglobin was successfully immobilized into the mesoporous molecular sieve MCFs, the immobilized enzyme was 50.27 mg g−1. The desirable adsorbate/adsorbent, Hb/MCFs, (mass ratio, m/m) for 3/50 was achieved by adjusting adsorption conditions. The obtained materials were characterized using various techniques such as the powder XRD, Fourier transform infrared spectroscopy, UV–Vis solid diffuse reflectance spectroscopy (SDRS), photoluminescence spectrum and N2 adsorption desorption study at 77 K. The results showed that hemoglobin had been immobilized successfully inside the channels of mesoporous MCFs. Scanning electron microscopy was used to confirm the average particle diameters of MCFs-Hb and (CH3-MCFs)-Hb. And the results showed that the average particle diameters of MCFs-Hb and (CH3-MCFs)-Hb were 2212 ± 50 nm and 2219 ± 50 nm, respectively. The adsorption process of MCFs/(CH3-MCFs) adsorbing hemoglobin enzyme was in accordance with pseudo-second-order kinetic equation. The process of MCFs/(CH3-MCFs) adsorbing hemoglobin enzyme belonged to an exothermic reaction which was a spontaneous reaction. Freundlich model described the adsorption of hemoglobin by MCFs/CH3-MCFs well. The desorption process of MCFs-Hb/(CH3-MCFs)-Hb composite was studied in the sodium hydroxide solution (0.1 mol L−1), and equilibrium was reached at 5 h. The desorption rates were 72.58% and 69.67%, respectively. The luminescence spectra showed that the obtained composites had the light-emitting properties.

Hemoglobin Immobilization on Multiporous Nanofibers of SnO₂ and Chitosan Composite for Hydrogen Peroxide Sensing.

A multiporous nanofiber (MPNFs) of SnO₂ and chitosan has been used for the immobilization of a redox protein, hemoglobin (Hb), onto the surface of glassy carbon electrode (GCE). The multiporous nanofiber of SnO₂ that has very high surface area is synthesized by using electrospinning technique through controlling the tin precursor concentration. Since the constructed MPNFs of SnO₂ exposes very high surface area, it increases the efficiency for biomolecule-loading. The morphology of fabricated electrodes is examined by SEM observation and the absorbance spectra of Hb/(MPNFs) of SnO₂ are studied by UV-Vis analysis. Cyclic Voltammetry and amperometry are employed to study and optimize the performance of the resulting fabricated electrode. After fabrication of the electrode with the Hb and MPNFs of SnO₂, a direct electron transfer between the protein's redox centre and the glassy carbon electrode was established. The modified electrode has showed a couple of redox peak located at -0.29 V and -0.18 V and found to be sensitive to H₂O₂. The fabricated electrode also exhibited an excellent electrocatalytic activity towards the reduction of H₂O₂. The catalysis currents increased linearly to the H₂O₂ concentration in a wide range of 5.0×10-6-1.5×10-4 M. Overall experimental results show that MPNFs of SnO₂ has a role towards the enhancement of the electroactivity of Hb at the electrode surface. Thus the MPNFs of SnO₂ is a very promising candidate for future biosensor applications.

Functionalized Black Phosphorus Nanocomposite for Biosensing

AbstractA functionalized black phosphorus (BP) nanocomposite was synthesized by non‐covalently combining ionic liquid (IL) and poly(diallyldimethylammonium chloride) (PDDA) with BP. The resultant nanocomposite exhibited good biocompatibility, favorable solubility, and enhanced electric conductivity to facilitate the immobilization of hemoglobin (Hb) and realize fast direct electron transfer with a rate constant (kET) of 15.36 s−1. The immobilized Hb displayed desirable electrocatalytic activity toward nitrite reduction in the range of 80 μM to 3.8 mM with a detection limit of 3.65 μM. The present work advances a new way to construct novel biofriendly BP‐based biosensors, biofuel cells, and bioelectronics with excellent sensitivity and stability.

Boron-doped Graphene quantum dots modified electrode for electrochemistry and electrocatalysis of hemoglobin

In this paper boron-doped graphene quantum dots (B-GQDs) decorated carbon ionic liquid electrode (CILE) was fabricated for the immobilization of hemoglobin (Hb), which was used as the electrochemical biosensor for electrocatalysis. Spectroscopic results demonstrated that the original structure of Hb was unchanged after mixed with B-GQDs, indicating the biocompatibility of B-GQDs. Electrochemical behaviors of Nafion/Hb/B-GQDs/CILE were investigated in pH 4.0 phosphate buffer solution and a couple of well-distinct symmetric redox peaks appeared on cyclic voltammograms. The results were also compared with GQDs and nitrogen-doped GQDs modified electrodes, which illustrated the enhanced effects of B-GQDs to electron communication and loading capability of Hb on the electrode surface due to its high conductivity and small size. Electrochemical kinetics were studied with the electrochemical parameters calculated. Electrocatalysis of Hb and B-GQDs modified electrode to various substrates such as trichloroacetic acid, sodium nitrite and hydrogen peroxide were carefully investigated. This new Hb modified electrode exhibited the advantages of large linear relationship, high sensitivity, low detection limit, good anti-interference ability and excellent stability, which found wide applications in food and drug samples detection.

Synthesis of a Poly-l-Lysine/Black Phosphorus Hybrid for Biosensors.

A simple, noncovalent modification strategy was proposed to synthesize poly-l-lysine-black phosphorus (pLL-BP) hybrid. BP nanoflakes were prepared with a water-phase exfoliation method. pLL can adhere to the surface of BP via hydrophobic interaction between butyl chains of pLL and the BP surface as well as the electrostatic interaction between the protonated amino groups on pLL and the negative charge on deprotonated PxOy groups remaining on BP. The as-synthesized pLL-BP hybrid turns out to be an ideal matrix for hemoglobin immobilization and direct electron transfer. Good conductivity and biocompatibility of BP maintain the native structure and the bioactivity of hemoglobin (Hb), facilitating the direct electron transfer between the electroactive center of Hb and electrode. The rate constant ( kET) for direct electron transfer of Hb@pLL-BP is calculated to be 11.24 s-1. The constructed Hb-pLL-BP based enzymatic electrochemical biosensor displays excellent catalytic activity toward the reduction of oxygen and hydrogen peroxide. The electrochemical response toward H2O2 exhibits a linear dependence on hydrogen peroxide concentration ranging between 10 μM and 700 μM. The results demonstrate that the pLL-BP hybrid can act as a biocompatible building block for the construction of novel biofuel cells, bioelectronics, and biosensors.

Facile fabrication of poly(glycidyl methacrylate)-b-polystyrene functional fibers under a shear field and immobilization of hemoglobin

PGMA-b-PS fibers were fabricated under a shear field for immobilization of bovine hemoglobin which has potential applications in blood substitutes.

Graphene dispersed cellulose microfibers composite for efficient immobilization of hemoglobin and selective biosensor for detection of hydrogen peroxide

In the present work, we have investigated the electrochemical behavior and electrocatalysis of hemoglobin (Hb) immobilized on a glassy carbon electrode (GCE) modified with a graphene-cellulose microfiber (GR–CMF) composite. The GR–CMF composite was characterized by scanning electron microscopy, elemental analysis, and Raman and Fourier transform infrared spectroscopy. Well-defined electrochemical redox characteristics of Hb were observed for Hb immobilized on a GR–CMF composite modified GCE, with a formal potential of −0.306V and a peak to peak separation of approximately 67mV. Due to the high biocompatibility of the GR–CMF composite, the electrochemical behavior of the Hb heme redox couple (FeII/FeIII) was enhanced for Hb immobilized on the GR–CMF composite when compared to Hb immobilized on pristine GR. The heterogeneous electron transfer constant (ks) was calculated as 6.17 s−1, and is higher than previously reported for Hb immobilized GR supports. The Hb immobilized GR–CMF composite modified electrode was used for the quantification of H2O2 under optimal conditions, and shows a wider linear amperometric response ranging from 0.05 to 926μM. The limit of detection of the biosensor was 0.01μM with the sensitivity of 0.49μAμM−1cm−2. The biosensor also showed high selectivity in the presence of the range of interfering compounds and exhibits good operational stability and practicality in the detection of H2O2.

1T‐Phase WS2 Protein‐Based Biosensor

Metallic 1T‐phase transition metal dichalcogenides have been recognized for their desirable properties like high surface‐to‐volume ratio, high conductivity, and capacitive behavior, making them outstanding for catalytic and sensing applications. Herein, a hydrogen peroxide (H2O2) biosensor is constructed by the immobilization of hemoglobin (Hb) on 1T‐phase WS2 (1T‐WS2) sheets, and entrapment by glutaraldehyde. 1T‐WS2 not only displays electrocatalytic activity toward the reduction of H2O2 but also provides a high surface‐to‐volume ratio and conductive platform for the immobilization of Hb and facilitation of its electron transfer to the electrode surface. The advantageous role of 1T‐phase WS2 is further demonstrated for the construction of a heme‐based H2O2 biosensor compared to its 1T‐phase MoS2, MoSe2, and WSe2 counterparts. Synergistic interactions between 1T‐WS2 and Hb result in a H2O2 biosensor with high analytical performance in terms of wide range, sensitivity, selectivity, reproducibility, repeatability, and stability. These findings have profound impact in the research fields of electrochemical sensing and biodiagnostics.

Bioelectrocatalysis of hydrogen peroxide based on immobilized hemoglobin onto glassy carbon electrode modified with magnetic poly(indole-co-thiophene) nanocomposite

On the basis of the emulsion polymerization process poly(indole-co-thiophene) supported Fe3O4 NPs nanocomposite was synthesized at room temperature. The structure and chemical composition of the nanocomposite were described by scanning electron microscopy, X-ray diffraction, and Fourier transform infrared spectroscopy. Due to its uniform pore structure, high surface areas and good biocompatibility, the chains of PIT@Fe3O4 nanocomposite are able to provide a suitable matrix for immobilization of biomolecule. Glassy carbon electrode modified with PIT@Fe3O4 nanocomposite was used as support for immobilization of hemoglobin (Hb) protein to fabricate an electrochemical biosensor. The electrochemical studies for the biosensor shows that it has excellent property for direct bioelectrocatalytic performances of hemoglobin, and could sense hydrogen peroxide at a low detection limit of 0.54μM and a linear range up to 350μM. The developed biosensor confirmed an excellent performance in the detection of hydrogen peroxide in environmental and food samples.

Modified fractal iron oxide magnetic nanostructure: A novel and high performance platform for redox protein immobilization, direct electrochemistry and bioelectrocatalysis application

A novel biosensing platform based on fractal-pattern of iron oxides magnetic nanostructures (FIOMNs) and mixed hemi/ad-micelle of sodium dodecyl sulfate (SDS) was designed for the magnetic immobilization of hemoglobin (Hb) at a screen printed carbon electrode (SPCE). The FIOMNs was successfully synthesized through hydrothermal approach and characterized by atomic force microscopy (AFM), scanning electron microscopy (SEM) and X-ray diffraction (XRD). In order to provide guidelines for the mixed hemi/ad-micelle formation, zeta-potential isotherms were investigated. The construction steps of the biosensor were evaluated by electrochemical impedance spectroscopy, cyclic voltammetry and Fourier transform infrared spectroscopy. Direct electron transfer of Hb incorporated into the biocomposite film was realized with a pair of quasi-reversible redox peak at the formal potential of −0.355V vs. Ag/AgCl attributing to heme Fe(III)/Fe(II) redox couple. The results suggested that synergistic functions regarding to the hyper-branched and multidirectional structure of FIOMNs and the dual interaction ability of mixed hemi/ad-micelle array of SDS molecules not only induce an effective electron transfer between the Hb and the underlying electrode (high heterogeneous electron transfer rate constant of 2.08s−1) but also provide powerful and special microenvironment for the adsorption of the redox proteins. Furthermore, the biosensor displayed an excellent performance to the electrocatalytic reduction of H2O2 with a detection limit of 0.48µM and Michaelis-Menten constant (Km) value of 44.2µM. The fabricated biosensor represented the features of sensitivity, disposable design, low sample volume, rapid and simple preparation step, and acceptable anti-interferences, which offer great perspectives for the screen-determination of H2O2 in real samples.

Direct electrochemistry of immobilized hemoglobin and sensing of bromate at a glassy carbon electrode modified with graphene and β-cyclodextrin

We describe the use of a nanocomposite consisting of graphene and β-cyclodextrin (β-CD) which was used to modify a glassy carbon electrode (GCE) to serve as a matrix for immobilization of hemoglobin (Hb). The composite was characterized by scanning electron microscopy, UV-vis and FTIR spectroscopy. The modified electrode displays an enhanced and well-defined quasi reversible peaks for the heme protein at a formal potential of −0.284 V (vs. Ag/AgCl). The direct electrochemistry of Hb is strongly enhanced at this modified electrode compared to electrodes not modified with graphene or β-CD. The heterogeneous electron transfer rate constant (Ks) is 3.18 ± 0.7 s−1 which indicates fast electron transfer. The biosensor exhibits excellent electrocatalytic activity towards the reduction of bromate, with a linear amperometric response in the 0.1 to 176.6 μM concentration range at a working voltage of −0.33 V. The sensitivity is 3.39 μA μM−1 cm−2, and the detection limit is 33 nM. The biosensor is fast, selective, well repeatable and reproducible, and therefore represents a viable platform for sensing bromate in aqueous samples.

Hemoglobin immobilized in exfoliated Co2Al LDH-graphene nanocomposite film: Direct electrochemistry and electrocatalysis toward trichloroacetic acid

A sandwich-like nanocomposite of exfoliated Co2Al layered double hydroxide (ELDH) and graphene (GR) was synthesized by chemically reducing in situ the assembly of ELDH and graphene oxide (GO) nanosheets. The morphology and structure of the product were characterized by scanning electron microscope (SEM), transmission electron microscope (TEM), X-ray diffraction (XRD) and atomic force microscopy (AFM). Then the ELDH-GR nanocomposite and chitosan (CTS) were employed for immobilization of hemoglobin (Hb) on a carbon ionic liquid electrode (CILE) to fabricate a trichloroacetic acid (TCA) biosensor. The UV-Vis, FTIR and fluorescence spectra showed that Hb kept its native structure and retained stable bioactivity in the composite material. Cyclic voltammetry (CV) displayed a pair of well-defined redox peaks with the formal peak potential (E0′) at −0.256V (vs. SCE), in pH 4.0 Britton–Robinson solution at the scan rate of 0.1V/s, implying the direct electron transfer of Hb–FeIII/FeII. The electron transfer coefficient (α) and the apparent heterogeneous electron transfer rate constant (ks) were calculated to be 0.492 and 1.202s−1, respectively. The Hb-modified electrode presented excellent electro-reduction activity toward TCA in the concentration range from 5 to 360mmol/L with the detection limit of 1.506mmol/L (3σ) by CV. For square wave voltammetry (SWV), the concentration range and the detection limit were from 2.5 to 360mmol/L and 0.82mmol/L, respectively. Therefore, the ELDH-GR composite has a promising potential for construction of biosensors.

Electrostatically Immobilized Hemoglobin on Silica-Coated Magnetic Nanoparticles for Simultaneous Determination of Dopamine, Uric Acid, and Folic Acid

This work describes the development of a novel biosensor for simultaneous determination of dopamine, uric acid, and folic acid by carbon paste electrode modified by hemoglobin which is electrostatically immobilized on silica-coated magnetic nanoparticles and multi-walled carbon nanotubes. Peroxidase properties of hemoglobin nominate this protein as an abundant and inexpensive alternative for costly peroxidase enzymes in dopamine biosensor fabrication. Silica-coated nanoparticles of Fe3O4 as a suitable support for hemoglobin immobilization can enhance the loading and stability of hemoglobin. Magnetic nanoparticles were synthesized via a simple co-precipitation method and then were coated with silica to immobilize hemoglobin electrostatically on them. All synthesis and modification steps were characterized by different techniques and then the conditions of hemoglobin immobilization on magnetic nanoparticles were optimized. Multi-walled carbon nanotubes were used as an additional modifier of carbon paste electrode because of their high electroactive surface area and interesting electrocatalytic properties. The modified carbon paste electrode provided a sensitive and stable biosensor for simultaneous determination of dopamine, uric acid, and folic acid whose performance is much better than those of many previously reported sensors. The detection limits were calculated to be about 12 nM, 14 nM and 18 nM and the linear range for determination of DA, UA and FA are 1–30.6 μM, 1–286 μM and 1–369 μM, for dopamine, uric acid, and folic acid, respectively. Finally, the applicability of the proposed biosensor was verified by dopamine evaluation in serum samples.

Immobilization of hemoglobin on functionalized multi-walled carbon nanotubes-poly-l-histidine-zinc oxide nanocomposites toward the detection of bromate and H2O2

A novel biocompatible sensing strategy has been developed based on functionalized multi-walled carbon nanotube (f-MWCNT), poly-l-histine (P-l-His), and ZnO nanocomposite film for the immobilization of hemoglobin (Hb). The direct electron transfer properties and bioelectrocatalytic activity of the Hb in f-MWCNT–P-l-His–ZnO composite film is further investigated. The apparent heterogeneous electron transfer rate constant (ks) of Hb confined to f-MWCNT–P-l-His–ZnO nanocomposite is found to be 5.16s−1 using Laviron's equation. Moreover, the surface coverage concentration (Γ) of the electroactive Hb in the f-MWCNT–P-l-His–ZnO film is estimated to be 1.88×10−9molcm−2. The fabricated electrochemical biosensor based on the immobilized Hb revealed a fast response time (<3s) with a wide linear range (4–18,000μM and 2–15,000μM) and detection limit (as low as 0.01μM and 0.30μM) for the electrocatalytic determination of a mediator-free H2O2 and bromate under optimal experimental conditions. The ca. apparent Michaelis–Menten constant is 0.14mM, which indicates that the Hb has a high affinity to H2O2. The high sensitivity, good reproducibility, and long-term stability of the proposed nanocomposite film indicates that it can serve as an electrode for the development of an amperometric H2O2 and bromate-based biosensor. The proposed third-generation biosensor was successfully applied to milk and urine samples for the detection of H2O2 and bromate.