Assessment of the efficiency and stability of enzymatic membrane reaction utilizing lipase covalently immobilized on a functionalized hybrid membrane.

Enzyme immobilization in membrane bioreactors has been considered as a practical approach to enhance the stability, reusability, and efficiency of enzymes. In this particular study, a new type of hybrid membrane reactor was created through the phase inversion method, utilizing hybrid of graphene oxide nanosheets (GON) and polyether sulfone (PES) in order to covalently immobilize the Candida rugosa lipase (CRL). The surface of hybrid membrane was initially modified by (3-Aminopropyl) triethoxysilane (APTES), before the use of glutaraldehyde (GLU), as a linker, through the imine bonds. The resulted enzymatic hybrid membrane reactors (EHMRs) were then thoroughly analyzed by using field-emission scanning electron microscopy (FE-SEM), contact angle goniometry, surface free energy analysis, X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, attenuated total reflection (ATR), and energy-dispersive X-ray (EDX) spectroscopy. The study also looked into the impact of factors such as initial CRL concentration, storage conditions, and immobilization time on the EHMR's performance and activity, which were subsequently optimized. The results demonstrated that the CRLs covalently immobilized on the EHMRs displayed enhanced pH and thermal stability compared to those physically immobilized or free. These covalently immobilized CRLs could maintain over 60% of their activity even after 6 reaction cycles spanning 50 days. EHMRs are valuable biocatalysts in developing various industrial, environmental, and analytical processes.

Carbon dot incorporated mesoporous silica nanoparticles for targeted cancer therapy and fluorescence imaging.

A new and efficient theranostic nanoplatform was developed via a green approach for targeted cancer therapy and fluorescence imaging, without the use of any anticancer chemotherapeutic drugs. Toward this aim, monodisperse and spherical mesoporous silica nanoparticles (MSNs) of approximately 50 nm diameter were first synthesized using the sol-gel method and loaded with hydrothermally synthesized anticancer carbon dots (CDs). The resulting MSNs-CDs were then functionalized with chitosan and targeted by an anti-MUC1 aptamer, using the glutaraldehyde cross-linker, and fully characterized by TEM, FE-SEM, EDS, FTIR, TGA, XRD, and BET analysis. Potent and selective anticancer activity was obtained against MCF-7 and MDA-MB-231 cancer cells with the maximum cell mortalities of 66.2 ± 1.97 and 71.8 ± 3%, respectively, after 48 h exposure with 100 µg mL-1 of the functionalized MSNs-CDs. The maximum mortality of 40.66 ± 1.3% of normal HUVEC cells was obtained under the same conditions. Based on the results of flowcytometry analysis, the apoptotic mediated cell death was recognized as the main anticancer mechanism of the MSNs-CDs. The fluorescence imaging of MCF-7 cancer cells was also studied after exposure with MSNs-CDs. The overall results indicated the high potential of the developed nanoplatform for targeted cancer theranostics.

Facile, rapid and efficient isolation of circulating tumor cells using aptamer-targeted magnetic nanoparticles integrated with a microfluidic device.

Facile and sensitive detection and isolation of circulating tumor cells (CTCs) was achieved using the aptamer-targeted magnetic nanoparticles (Apt-MNPs) in conjugation with a microfluidic device. Apt-MNPs were developed by the covalent attachment of anti-MUC1 aptamer to the silica-coated magnetic nanoparticles via the glutaraldehyde linkers. Apt-MNPs displayed high stability and functionality after 6 months of storage at 4 °C. The specific microfluidic device consisting of mixing, sorting and separation modules was fabricated through conventional photo- and soft-lithography by using polydimethylsiloxane. The capture efficiency of Apt-MNPs was first studied in vitro on MCF-7 and MDA-MB-231 cancer cell lines in the bulk and microfluidic platforms. The cell capture yields of more than 91% were obtained at the optimum condition after 60 minutes of exposure to 50 µg mL-1 Apt-MNPs with 10 to 106 cancer cells in different media. CTCs were also isolated efficiently from the blood samples of breast cancer patients and successfully propagated in vitro. The isolated CTCs were further characterized using immunofluorescence staining. The overall results indicated the high potential of the present method for the detection and capture of CTCs.

Controlled drug delivery and cell adhesion for bone tissue regeneration by Keplerate polyoxometalate (Mo132)/metronidazole/PMMA scaffolds.

The aim of this study is to fabricate a new scaffold appropriate for tissue regeneration with antimicrobial activity and ability of controlled drug delivery. In this regard, scaffold nanofibers were produced using poly (methyl methacrylate) (PMMA), Mo132 as a Keplerate polyoxometalate and metronidazole. The final scaffolds, obtained by electrospinning, represent the intrinsic features including exceptional doubling tensile strength, high hydrophilicity (126 ± 5.2° to 83.9 ± 3.2° for contact angle and 14.18 ± 0.62% to 35.62 ± 0.24% for water uptake), proper bioactivity and cell adhesion. Moreover, the addition of Mo132 and metronidazole enhances the biodegradation rate of resulted scaffolds compared to the pure PMMA membrane. The controlled release of metronidazole over 14 days efficiently inhibits the colonization of anaerobic microorganisms. Overall, the results demonstrate high potential of Mo132 and metronidazole-loaded PMMA scaffold for guided bone regeneration/guided tissue regeneration.

Fluorescence resonance energy transfer monitoring of pH-responsive doxorubicin release from carbon dots/aptamer functionalized magnetic mesoporous silica.

Aim: To develop a novel theranostic nanoplatform for simultaneous fluorescent monitoring and stimuli-triggered drug delivery. Materials & methods: Different microscopic and spectroscopic techniques were used for the characterization of nanocarriers. MCF-7 and human umbilical vein endothelial cell lines were cultured and treated with different doses of doxorubicin-loaded nanocarriers. The cell viability and drug release were studied using MTT assay and fluorescence microscopy. Results: Biocompatible and mono-disperse nanocarriers represent hollow and mesoporous structures with the calculated surface area of 552.83 m2.g-1, high magnetic activity (12.6 emu.g-1), appropriate colloidal stability and high drug loading capacity (up to 61%). Conclusion: Taxane-based carbon dots act as the pH-responsive gatekeepers for the controlled release of doxorubicin into cancer cells and provide a fluorescence resonance energy transfer system for real-time monitoring of drug delivery.

PEGAylated graphene oxide/superparamagnetic nanocomposite as a high-efficiency loading nanocarrier for controlled delivery of methotrexate.

Polymer-coated nanocarriers play an important role in targeted drug delivery. The use of polymers such as polyethylene glycol increases stability, biocompatibility, and blood circulation time of the drug, and may consequently improve the success of drug delivery. In the present work, a simple approach has been reported for synthesizing polyethylene glycol bis amin (PEGA) functionalized graphene oxide/iron oxide nanocomposite as a remarkable unit for loading drugs. The biomedical applications of the synthesized nanocomposite were investigated by immobilizing methotrexate (MTX), as an anticancer drug. The structural and morphological characteristics and the successful synthesis of the nanocomposite were evaluated by different charachterization techniques. The cytotoxicity assay of the nanocarrier showed higher toxicity against HeLa and MCF-7 cell lines, compared to free MTX. The drug release experiments in acidic and physiological conditions suggested the first order kinetics model for the release of MTX from the nanocomposite. Furthermore, the agglutination, complement activation, and coagulation time experiments demonstrated the blood compatibility of the synthesized nanocarrier.

Electrochemiluminescence detection of human breast cancer cells using aptamer modified bipolar electrode mounted into 3D printed microchannel.

In the present manuscript, a closed bipolar electrode system integrated with electrochemiluminescence (ECL) detection has been introduced for sensitive diagnosis of human breast cancer cells (MCF-7). For sensitive and selective detection, the anodic pole of the bipolar electrode was modified with the AS1411 aptamer, a specific aptamer for the nucleolin, and treated by the secondary aptamer modified gold nanoparticles. The electrochemiluminescence of luminol was followed in the presence of hydrogen peroxide on the anode pole of bipolar electrode (BPE) as an analytical signal. Moreover, 3D printed microchannels were used for the fabrication of BPE systems to minimize the required amounts of sample. The present aptasensor offers low cost, sensitive and selective cancer cell detection with two acceptable linear ranges. The first linear section appears within 10-100 cells and the latter is found to be within 100-700 cells. The limit of detection was about 10 cells.

Self-recognition of the racemic ligand in the formation of homochiral dinuclear V(V) complex: In vitro anticancer activity, DNA and HSA interaction.

The reaction of a racemic mixture of Schiff base tridentate ligand with vanadium(V) affords homochiral vanadium complex, (VO(R-L))2O and (VO(S-L))2O due to ligand "self-recognition" process. The formation of homochiral vanadium complex was confirmed by 1H NMR, 13C NMR and X-ray diffraction. The HSA- and DNA-binding of the resultant complex is assessed by absorption, fluorescence and circular dichroism (CD) spectroscopy methods. Based on the results, the HSA- and DNA-binding constant, Kb, were found to be 8.0 × 104 and 1.9 × 105 M-1, respectively. Interestingly, in vitro cytotoxicity assay revealed the potent anticancer activity of this complex on two prevalent cancer cell lines of MCF-7 (IC50 value of 14 µM) and HeLa (IC50 value of 36 µM), with considerably low toxicity on normal human fibroblast cells. The maximum cell mortality of 12.3% obtained after 48 h incubation of fibroblast cells with 100 µM of the complex. Additionally, the specific DNA- and HSA-binding was also shown using molecular docking method. The synthesized complex displayed high potential for biomedical applications especially for development of novel and efficient anticancer agents.

Synthesis, characterization and separation of chiral and achiral diastereomers of Schiff base Pd(II) complex: A comparative study of their DNA- and HSA-binding.

A racemic mixture of a new chiral Schiff base ligand (HL: R/S-(1-phenylethylimino)methylnaphtalen-2-ol) has been utilized to prepare Pd(II) complex. Crystallization technique has been employed to separate diastereomeric pairs of Pd(II) complex: (mesoPdL2) and (racPdL2) that in this paper are known as PdL2(1) and PdL2(2), respectively. The synthesized complexes have been characterized by means of elemental analysis (CHN), FT-IR, (1)H and (13)C NMR spectroscopies. Moreover, PdL2(1) has been structurally characterized by single-crystal X-ray diffraction. The geometry around the metal center is square-planar. The interaction of two diastereomers of Pd(II) complex with FS-DNA has been explored, using UV-vis spectroscopy, fluorescence quenching, chemometrics and viscosity measurement methods. The PdL2(1) exhibited higher binding constant, about 10-fold, (1.0×10(6)M(-1)) as compared to PdL2(2) (1.51.5×10(5)M(-1)). Moreover, the human serum albumin (HSA) binding ability has been monitored by absorption, quenching of tryptophan fluorescence emission and circular dichroism (CD) studies. The slight difference is observed between HSA binding affinity with the complexes: PdL2(1) (6.2×10(4)M(-1)) and PdL2(2) (3.3×10(4)M(-1)). Also, the thermodynamic parameters were determined at three different temperatures (298, 308 and 318K). In this study, molecular docking was also carried out to confirm and illustrate the specific DNA- and HSA-binding of the Pd(II) complexes. In the PdL2(1)-HSA system a T-shaped π-π interaction with PHE206 was observed. While in the PdL2(2)-HSA system there are a hydrogen bond, a π-cation and two T-shaped π-π interactions with ASB324, LYS212 and PHE228, respectively. The groove binding mode of DNA interaction has been proposed for both diastereomers.

Optimization of the basal medium for improving production and secretion of taxanes from suspension cell culture of Taxus baccata L.

BACKGROUND AND PURPOSE OF THE STUDY: Taxol is one of the most effective anticancer drugs that isolated from Taxus sp. due to the slow growth of Taxus trees and low concentration of Taxol in the tissues, the biotechnological approaches especially plant cell culture have been considered to produce Taxol in commercial scale. METHODS: We investigated the effects of basal medium type used in culture media on production of Taxol and other taxane compounds from cell suspension culture of T. baccata L. Briefly, five commonly basal media including Gamborg, Murashige and Skoog, Woody Plant, Schenk and Hildebrandt, and Driver and Kuniyuki medium were used for preparing separate suspension culture media. The intra- and extra-cellular yields of taxanes were analyzed by using HPLC after 21 days period of culturing. RESULTS: The yields of taxanes were significantly different for the cultures prepared by different basal media. Moreover, the effects of basal medium on the yield of products differed for varius taxane compounds. Maximum yields of Baccatin III (10.03 mgl-1) and 10-deacetyl baccatin III (4.2 mgl-1) were achieved from the DKW basal media, but the yield of Taxol was maximum (16.58 mgl-1) in the WPM basal media. Furthermore, the secretion of taxanes from the cells into medium was also considerably affected by the type of basal medium. The maximum extra-cellular yield of Taxol (7.81 mgl-1), Baccatin III (5.0 mgl-1), and 10-deacetyl baccatin III (1.45 mgl-1) were also obtained by using DKW basal medium that were significantly higher than those obtained from other culture media.

Enhanced taxane production and secretion from Taxus baccata cell culture by adding dimethylsulfoxide.

The ability of an aprotic solvent, dimethylsulfoxide (DMSO), to induce taxane synthesis and release from cell suspension culture of Taxus baccata was examined. The results showed that applying DMSO in optimal conditions not only led to enhancement in taxane excretion from the cells but also led to improvement in taxol synthesis. Maximum yields for taxol [3.34 mg/g dry cell weight (DCW)] were achieved by adding 5% DMSO to the culture at the late-exponential phase of cell growth (day 14) and culturing for 21 days, which was 2.26-fold of that for the control (1.48 mg/g DCW). However, adding 5% DMSO did not affect the yield of 10-deacetyl baccatin III and baccatin III. This condition also resulted in maximum extracellular taxane (4.86 mg/L); this value was 2.82-fold higher than that in the control (1.72 mg/L). We demonstrated that the late-exponential phase of cell growth could be the best time to add elicitor for maximizing the yield of taxol. Adding DMSO at earlier times (days 1 and 7) or in other concentrations (1% and 3%) had negative effects on taxane synthesis. Overall results suggest that DMSO has good potential to enhance synthesis and release taxol from cell suspension culture of T. baccata L.

Encapsulated activated charcoal as a potent agent for improving taxane synthesis and recovery from cultures.

The ability of a potent hydrophobic adsorbent, AC (activated charcoal), to induce total taxane synthesis and to improve the recovery of the products from suspension culture of Taxus baccata L. was examined. The results strongly showed that the application of encapsulated AC as an adsorbent not only led to improvement of in situ recover but also led to enhancement in taxane synthesis within the cells as well as having no inhibitory effects on the cell growth. The maximum yields for 10-deacetyl baccatin III, baccatin III and taxol were achieved by adding 1 g.l-1 adsorbent as alginate beads to the culture and culturing for 21 days and these were 3.5-, 4- and 2-fold respectively of that for the control. We demonstrated that the late exponential phase of the cell growth (day 14) could be the best time to add the adsorbent for maximizing the yield of taxol. Adding 1 g.l-1 AC resulted in maximum extracellular taxol (5.584 mg.l-1); this value was 2.19-fold higher than that in the control. According to the present results, AC is a powerful adsorbent that efficiently adsorbs the taxane compounds from a medium. We suggested that encapsulated AC could be used as a promising adsorbent agent for large-scale taxane production when it is added in combination with proper elicitors.