Biomedical uses of Enzymes Immobilized by Nanoparticles

Fe3O4 and Fe3O4@SiO2 magnetic nanoparticles (MNPs) have recently become important in biomedical applications; however, influences of these MNPs to cells are still not very clear. Bare Fe3O4 and Fe3O4@SiO2 MNPs should be noticed because any surface modification may be removed from them when they enter into cells or in cells. In this work, in order to avoid too much surface residues from the precursors, coprecipitation method is adopted to synthesize bare Fe3O4 MNPs, while Stober process is performed to synthesize bare Fe3O4@SiO2 MNPs. The characterization of MNPs is indentified by X-ray Diffraction (XRD), Transmission Electron Microscopy (TEM), X-ray Photoelectron Spectroscopy (XPS), X-ray Absorption Spectroscopy (XAS) and Superconducting Quantum Interference Device Magnetometer (SQUID). These results show that as-prepared Fe3O4 MNPs primarily contains crystalline Fe3O4 phase, while the deposited SiO2 on Fe3O4 MNPs is amorphous. A549 lung cancer cells are used as model cells for MNPs treatment, and the cell viability is measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results show that mitochondrial reductase activity in cells is reduced by treating Fe3O4 MNPs and Fe3O4@SiO2 MNPs to A549 cells for 36 hr. Instead of traditional biochemical methods, synchrotron radiation infrared-ray (SRIR) spectra and synchrotron radiation infrared-ray microscopy (SRIRM) with high spatial resolution 10μm are carried out to measure the change of chemical components and chemical composition distribution in cells. These results exhibit that DNA structures in cells are indirectly affected by Fe3O4 MNPs and Fe3O4@SiO2 MNPs, and the concentration of DNA becomes less with MNPs concentration and treatment time while no protein and lipid changes are observed, but the lipid/protein ratio is MNPs-concentration-dependent and treatment-time-dependent and it is observed that the amount of lipids is relatively larger at far-nucleus regions while that of proteins is relative larger at and around the nucleus region.

Introduction: Magnetic nanoparticle (MNP) hyperthermia is a promising cancer treatment approach. It is based on the evidence that by injecting MNPs such as Fe3O4 in the tumor and subjecting them to an alternating magnetic field, they release heat, generating temperatures up to 42°C that can kill cancer cells by apoptosis, usually with lowest damage to normal tissue. In previous work the temperature distribution of an ordinary tumor over the different sizes of Fe3O4 MNPs with uniform dispersion in tumor was performed. The objective of this work is to evaluate the thermal distribution of MNPs in the tumor tissue using random and array geometries due to dispersion states of the inclusions. Materials and Methods: The bio heat transfer equation (BHTE) formulated by Pennes, describes thermal processes in the human body. Since finding the exact solution of BHTE for a complex system like tumor tissue is a challenging issue, so finite element method in the COMSOL Multiphysics software is used to capture more details in predicting the temperature distribution in tumor tissue during the hyperthermia therapy. Producing the random and array numbers for distribution of the MNPs into the tumor tissue was carried out with MATLAB software. Fe3O4 MNPs with the size of 100 nm with different distributions in the tumor were simulated and the temperature distribution of the tumor and normal tissue was calculated, taking into account the thermal conductivity, density, and heat capacity. Results: The distribution of temperature in the tumor depends on Fe3O4 MNPs distribution. Uniform distribution of MNPs has appropriate distribution in the tumor and normal tissue around it. And also the agglomeration of MNPs in the tumor tissue leads to non-uniformity of the temperature distribution of the inclusions. Conclusion: In this research, the temperature distribution of an ordinary tumor over the different distributions (random and array) of Fe3O4 MNPs was investigated via the finite element method. The bio heat transfer equation was used to calculate the thermal processes in the tumor and normal tissues around it. It should be taken into account the distribution states of the MNPs into the tumor tissue and its subsequently effects on the thermal distribution. Finally, results showed that uniform distribution of the magnetic nanoparticles into the tumor tissue exhibited more appropriated results.

Comparison of MnO2 modified and unmodified magnetic Fe3O4 nanoparticle adsorbents and their potential to remove iron and manganese from aqueous media

Multifunctional magnetic composite nanoparticles (NPs) with antibiotics have demonstrated symbiotic effects because of their promising antimicrobial properties. The antimicrobial agent reduces side effects and dosage, and increases drug delivery efficiency. In this study, SiO2 coated over Fe3O4 magnetic nanoparticles (MNPs) were prepared by a solvothermal method. The MNPs were characterized by using X-ray diffraction (XRD), transmission electron microscopy (TEM), ultraviolet-visible spectroscopy (UV-vis), and Fourier transform infrared spectroscopy (FTIR). The antimicrobial tests were carried out using the disk diffusion method. The electrochemical sensing was investigated by cyclic voltammetry with varying As(III) concentrations from 1–10 ppb. The microstructural results showed the formation of spherical-shaped Fe3O4@SiO2 MNPs with 15–30 nm diameters. UV-vis results showed that Fe3O4 NPs promote visible light absorption of Fe3O4@SiO2 MNPs because of well-structured and unvarying shell thickness which is beneficial for the absorption of organic dyes. With an increase in the concentration of As(III), there was a shift in potential and an increase in oxidation peak current, showing the electrocatalytic capacity of the modified electrode. The SiO2 deposited on Fe3O4 displayed an admirable microbial operation. These Fe3O4@SiO2 MNPs are easily absorbed by cells and have the potential to influence bacterial cells both within and outside of the cell membrane, making them an intriguing candidate for use in a variety of biological applications in the future.

Magnetic graphite carbon spheres (MGCSs) with well-distributed Fe3O4 magnetic nanoparticles were synthesized via the following steps. The colloidal carbon spheres (CCSs) with uniformly dispersed Fe(II) and large numbers of hydroxyl groups were synthesized via synchronous hydrothermal reaction of glucose and ferrous gluconate. The CCSs were then converted to MGCSs via consequent thermal treatment. In addition, the hydroxyl groups of the as-prepared CCSs were also utilized to adsorb Ag+ ions or condensate with Ti–OH. Following a thermal treatment, composite MGCS@Ag or MGCS@TiO2 microspheres were fabricated. The results of SEM, TEM and XRD revealed that MGCSs with an average diameter of 1 μm were synthesized; and magnetic Fe3O4 nanoparticles with diameters from 20 to 25 nm were uniformly distributed in MGCSs, which indicated that Fe(II) in the CCSs not only changed into magnetic Fe3O4 nanoparticles, but also worked as a catalyst for the graphitization of amorphous carbon during the thermal treatment. Such a strategy gave at least two advantages. First, the protective effect on the magnetic nanoparticles and mechanical strength of the graphite carbon may improve the capability and feasibility of practical applications. Synchronously, the MGCSs not only have well adsorbing properties as carbon materials, but also possess unusual adsorbing behaviours for heavy metal ions and noble metal ions, which have significant potential applications in the treatment of polluted water and the recovery of noble metals. Second, the synthesis of composite MGCSs may further expand the scope of the applications. For example, the MGCS@Ag and MGCS@TiO2 microspheres prepared in this effort exhibited excellent antibacterial activity and selective enrichment of phosphopeptides, respectively.

Green synthesis of chitosan magnetic nanoparticles and their application with poly-aldehyde kefiran cross-linker to immobilize pectinase enzyme

Objective To investigate the effect of targeted Fe3 O4 magnetic multidrug resistance (MDR) 1 siRNA nanoparticles on the rat BT325 glioblastoma multiform model.Methods 20 BT325 glioblastoma multiform rat models were build up.They were divided into 2 groups randomly:the Fe3 O4 magnetic MDR1 siRNA nanoparticles group and control group.After infusing of the nanoparticles and targeting by the outside magnetic material,the effects were evaluated by the tumor grows and MRI findings.The HE immunocytology of tumor and the FCM were also performed to check the changing of the tumor.Results The rat BT325 glioblastoma multiform model was made successfully.The tumor grew slowly after infusion the Fe3 O4 magnetic MDR1 siRNA nanoparticles and the weight of the rat became less on day 2.The image of MRI showed high signal on T2WI.HE immunocytology presented Fe particles expression.The FCM demonstrated the apoptosis appeared combined the control group.Conclusions Targeted Fe3 O4 magnetic MDR1 siRNA nanoparticles on the rat BT325 glioblastoma multiform model can inhibit tumor growth in vivo and it will increase chemotherapic sensitivity for glioblastoma multidrug resistance. Key words: Animal model ; Nanoparticles; Multidrug resistance ; Glioma

A type of illness known as cardiovascular disease affects the heart or blood arteries. The flow of blood to the heart, brain, or body is reduced due to thrombosis (blood clotting). Streptokinase (SK) is an extracellular enzyme that converts plasminogen to plasmin, as a medication in thrombolytic treatment. The current study was focused on streptokinase production and enhanced activity using magnetic nanoparticles. Nutrient agar media and liquid state fermentation at pre-optimized conditions was used. The proteolytic activity of the enzyme was determined by using the casein digestion method and then Biuret test was performed for protein estimation. The product of bacterial culture was extracted, and the partial purification of streptokinase was done by ammonium sulfate precipitation, dialysis and then further chromatographic techniques and gel filtration were applied. Immobilization of streptokinase was done by using Fe3O4 magnetic nanoparticles to enhance its activity and protein yield. The immobilization of SK on magnetic nanoparticles was characterized by UV-Vis spectroscopy and Zetasizer. Fe3O4 magnetic nanoparticle showed absorption peak at 224.4 nm and size of 229.4 nm. When compared to the total activity of gel chromatography (5.10 U/ml), immobilization increased the total activity of SK to 7.106 U/ml. In this way enzyme activity of immobilized streptokinase increased by 1.39-fold. Immobilized enzyme was used in in vivo studies and then compared with native one. The activity analysis of immobilized streptokinase was compared with the native one. Immobilized streptokinase exhibited more thrombolytic activity in rabbits than the native form, as demonstrated by in vivo study. Keywords: Streptokinase; Immobilized enzyme; Nanoparticle; Chromatography; Streptococcus pyogenes; Thrombosis

Magnetic nanoparticles towards efficient adsorption of gram positive and gram negative bacteria: An investigation of adsorption parameters and interaction mechanism

One of the remarkable features of bacterial species is their capacity for rapid growth when the appropriate environmental condition for growth is provided. Some bacteria, during their growth period, encounter stress factors in their natural environments, such as limitation in growth bioavailability, heat shock, heavy metal, etc. One stress factor not studied is the effect of magnetic Fe3O4 nanoparticles on bacterial growth rate. The effect of magnetic Fe3O4 nanoparticles on the protein profiles of genetically engineered bacterial strain Pseudomonas aeruginosa (PTSOX4), a strain with biological desulfurization characteristic, was investigated. The magnetic Fe3O4 nanoparticles were synthesized using co-sedimentation method, and their morphology was observed by scanning electron microscopy (SEM). The topography of magnetic Fe3O4 nanoparticles was detected by X-ray diffraction, and the average nanoparticle size measured was 40 to 50 nm. The bacterial cells were coated with magnetic nanoparticles, and the SEM electrographs of the bacterial cells indicated that the nanoparticles were uniformly coated on the cell surface. Proteins from both uncoated and coated bacterial cells were extracted by sonication and subjected to two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis. Some novel protein bands appeared in the protein profiles of coated bacterial cells; however, some protein bands disappeared. The two-dimensional gel electrophoresis results highlighted the presence of two different polypeptide groups, with molecular weights of 30 to 56 kDa and 56 to 65 kDa.

Rhodococcus erythropolis R1 is a capable strain in bioconversion of dibenzothiophene (DBT) to 2-hydroxybiphenyl (2-HBP) in oil model. In order to prevent the contamination of biodesulfurization (BDS) products by free cells, microbial cells were immobilized using different materials such as magnetic Fe3O4 nanoparticles (NPs). In this study, magnetic NPs were produced by two different procedures and their characteristics were determined via transmission electron microscopy (TEM) and X-ray diffraction (XRD). Also, binding of NPs on the cell surface was studied and better NPs were used for cells immobilization. Both NPs were crystallized and less than 10nm. The BDS by immobilized cells was carried out in biphasic system, and media conditions were optimized statistically by response surface methodology (RSM). The DBT concentration, temperature and interaction between them had statistically significant effects on 2-HBP production by nanomagnet immobilized cells. The optimum DBT concentration, temperature and pH for 2-HBP production by immobilized R. erythropolis R1 were obtained at 6.76mM, 29.63 °C and 6.84 respectively by HPLC analysis.

Biopolyester from ricinoleic acid: Synthesis, characterization and its use as biopolymeric matrix for magnetic nanocomposites

The magnetic nanoparticles (MNPs) of oxide with metal ions (MeIs-O) encapsulated with biological membranes or decorated with different organic shells are widely used in nano-biotechnology and nanomedicine, either as drug carriers in diagnosis and therapeutic biomedical fields or as a result of their specific magnetic effects on biological tissues. The MNP applications in pharma are closely related with their coating, shape, surface charge, core, and hydrodynamic size. The main characteristic of MNPs is the possibility to manipulate them with an external magnetic field (e-MF), which strongly depends on the magnetic structure of nanoparticles (NPs), multi-domain structure, in the case of NPs having sizes greater than approximately 30 nm or single domain in the case of NPs of sizes generally below 30 nm (depending on the type of NPs). A special structure of MNPs is the superparamagnetic (SPM) state when the magnetization of single-domain NPs is not stable and fluctuates along the easy magnetization axis under the action of thermal activation. This effect generally occurs when NPs have sizes smaller than approximately 15 nm. Depending on the targeted application in biotechnology/nanomedicine, only the NPs having the magnetic characteristics suitable for the type of application envisaged should be considered, in order to obtain the expected effect. In this chapter, we present the specific magnetic properties of MNPs depending on their size, the physical principle of handling NPs with an e-MF, magnetization of NPs, biocompatible MNPs (BC-MNPs) with salicylic acid (SA) and other organic coating, vascular nanoblocking of tumors, in vivo results using the chick embryo chorioallantoic membrane (CAM) model, and cytotoxicity of SA-MNPs.

Papain is widely used in food, drug, and bioactive peptide production and must be immobilized onto carriers with biocompatibility. Dialdehyde starch (DAS) can be a good biocompatible cross‐linker according to its active aldehyde groups. In the present study, the magnetic nanoparticles dialdehyde starch (MDASN), synthesized by DAS and Fe3O4 magnetic nanoparticles (Fe‐MNP), are successfully used to immobilize papain to improve the enzymic activity. The structure and morphology of DAS, MDASN, and immobilized papain onto magnetic dialdehyde starch nanoparticles (papain‐MDASN) are characterized detailly. The morphology of DAS is like a flat ball, and that of Fe‐MNP and papain‐MDASN are spherical and clumpy. The particle size of Fe‐MNP and papain‐MDASN are small, resulting in a large surface electrostatic effect and partial agglomeration. Enzymic activity studies of papain‐MDASN exhibit that the immobilized papain on MDASN represents better temperature resistance, alkaline resistance, thermal stability, and reusability, and its activity recovery is up to 68.21%. Papain onto magnetic dialdehyde starch nanoparticles (MDASN) may enhance its potential application in production processes.

The present study synthesizes a novel adsorbent by coating Fe3O4 magnetic nanoparticles with amino functionalized mesoporous silica. The FTIR spectrums indicate that silica has been successfully coated on the surface of Fe3O4 and 3-aminopropyl tri methoxysilane compound have been grafted to the surface of silica-coated Fe3O4. The XRD analysis shows the presence of magnetite phase with cubic spinel as a highly crystalline structure, before and after silica coating. The study also investigates the potentials of amino functionalized silica-coated Fe3O4 magnetic nanoparticles for extraction of Pb2+ and Cd2+ cations from aqueous solutions, where it has used flame atomic absorption spectrometry to determine ion concentration in both recovery and sample solutions. The optimum conditions of removal of Pb2+ and Cd2+ ions turn out to be pH= 4-8 with a stirring time of 20 minutes. The minimum amount of 3M nitric acid to strip ions from functionalized magnetic nanoparticles is 10 mL. The experimental data show the adsorption isotherms have been well described by Langmuir isotherm model, with the maximum capacity of the adsorbent being 1000.0 (± 1.4) μg, 454.5 (± 1.6) μg of Pb2+, and Cd2+ per each mg of functionalized magnetic nanoparticles, respectively. Finally, the proposed adsorbent is successfully applied to remove Pb2+ and Cd2+ ions in wastewater samples.