Impact of Lipid Composition on Vesicle Protein Adsorption: A BSA Case Study.

Investigating the interaction between liposomes and proteins is of paramount importance in the development of liposomal formulations with real potential for bench-to-bedside transfer. Upon entering the body, proteins are immediately adsorbed on the liposomal surface, changing the nanovehicles' biological identity, which has a significant impact on their biodistribution and pharmacokinetics and ultimately on their therapeutic effect. Albumin is the most abundant plasma protein and thus usually adsorbs immediately on the liposomal surface. We herein report a comprehensive investigation on the adsorption of model protein bovine serum albumin (BSA) onto liposomal vesicles containing the zwitterionic lipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), in combination with either cholesterol (CHOL) or the cationic lipid 1,2-dioleoyl-3-trimethylammoniumpropane (DOTAP). While many studies regarding protein adsorption on the surface of liposomes with different compositions have been performed, to the best of our knowledge, the differential responses of CHOL and DOTAP upon albumin adsorption on vesicles have not yet been investigated. UV-vis spectroscopy and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a strong influence of the phospholipid membrane composition on protein adsorption. Hence, it was found that DOTAP-containing vesicles adsorb proteins more robustly but also aggregate in the presence of BSA, as confirmed by DLS and TEM. Separation of liposome-protein complexes from unadsorbed proteins performed by means of centrifugation and size exclusion chromatography (SEC) was also investigated. Our results show that neither method can be regarded as a golden experimental setup to study the protein corona of liposomes. Yet, SEC proved to be more successful in the separation of unbound proteins, although the amount of lipid loss upon liposome elution was higher than expected. In addition, coarse-grained molecular dynamics simulations were employed to ascertain key membrane parameters, such as the membrane thickness and area per lipid. Overall, this study highlights the importance of surface charge and membrane fluidity in influencing the extent of protein adsorption. We hope that our investigation will be a valuable contribution to better understanding protein-vesicle interactions for improved nanocarrier design.

Chitosan nanofibers encapsulating copper oxide nanoparticles: A new approach towards multifunctional ecological membranes with high antimicrobial and antioxidant efficiency.

This paper focuses on the preparation of chitosan-based nanofibers embedding copper oxide nanoparticles to create multifunctional materials that meet the demands of contemporary applications. To this end, a mixture of chitosan, quaternized chitosan and poly (ethylene glycol) was used as polymeric matrix, considering their own contribution to the final material's properties and their ability to stabilize the copper oxide nanoparticles. An exhaustive investigation of the nanofibers was done in order to assess their composition and morphology (FTIR, 1H NMR, WXRD, TGA, SEM, TEM, POM, UV-vis) and to study their mechanical, antimicrobial and antioxidant properties, air and water permeability and ability for air filtration. It was shown that the copper oxide nanoparticles were anchored into the polymeric matrix via strong hydrogen bonding and electrostatic interactions, which induced the improvement of the mechanical properties and antioxidant activity. The copper oxide nanoparticles favored the thinning of the fibers during electrospinning process and improved the antibacterial activity and dust filtration capacity. Besides, the fibers displayed air permeability and vapor water transmission rate similar to synthetic nanofibers, while being biodegradable. All these performances recommend the new materials for developing antibacterial eco-materials with good breathability to be used as hygienic textiles, masks, or air filters.

Biodegradable trimethyl chitosan nanofiber mats by electrospinning as bioabsorbable dressings for wound closure and healing.

The paper aimed to prepare quaternary chitosan-based nanofibers as bioabsorbable wound dressings. To this aim, fully biodegradable chitosan/N,N,N-trimethyl chitosan (TMC) nanofibers were designed and prepared via electrospinning, using poly(ethylene glycol) as sacrificial additive. The new biomaterials were structurally and morphologically characterized by FTIR and NMR spectroscopy, thermogravimetric analysis, X-ray diffraction and scanning electron microscopy, and their properties required for wound dressings application were investigated and discussed in detail. Thus, the nanofiber behavior was investigated by swelling, dynamic vapor sorption, and in vitro biodegradation in media mimicking the wound exudate. The mechanical properties were analysed from the stress-strain curves, the bioadhesivity from the texture analysis and the mucoadhesivity from the Zeta potential and transmittance measurements. The antimicrobial activity was assessed against S. aureus and E. coli strains, and the biocompatibility was tested in vitro on normal human dermal fibroblasts, and in vivo on rats. The application of the fiber mats with the best balance of properties as dressings on deep burn wound models in rats showed wound closure and active healing, with fully restoration of epithelia. It was concluded that the combination of chitosan with TMC into nanofibers provides new potential bioabsorbable wound dressing, opening new perspectives in regenerative medicine.

Drug delivery based on a supramolecular chemistry approach by using chitosan hydrogels.

Microbial infections are a serious healthcare related problem, causing several complications and even death. That is why, the development of new drug delivery systems with prolonged effect represents an interesting research topic. This study presents the synthesis and characterization of new hydrogels based on chitosan and three halogenated monoaldehydes. Further, the hydrogels were used as excipients for the development of drug delivery systems (DDS) by the incorporation of fluconazole, an antifungal drug. The systems were structurally characterized by Fourier Transformed Infrared Spectroscopy and Nuclear Magnetic Resonance, both methods revealing the formation of the imine linkages between chitosan and the aldehydes. The samples presented a high degree of ordering at supramolecular level, as demonstrated by WXRD and POM and a good water-uptake, reaching a maximum of 1.6 g/g. The obtained systems were biodegradable, loosing between 38 and 49 % from their initial mass in the presence of lysozyme in 21 days. The ability to release the antifungal drug in a sustained manner for seven days, along with the high values of the inhibition zone diameter, reaching a maximum of 64 mm against Candida parapsilosis for the chlorine containing sample, recommend these systems as promising materials for bioapplications.

Mesoporous chitosan nanofibers loaded with norfloxacin and coated with phenylboronic acid perform as bioabsorbable active dressings to accelerate the healing of burn wounds.

The paper reports new chitosan-based nanofibers, designed to address the healing of burn wounds. To this aim, mesoporous chitosan fiber mats were prepared by electrospinning using poly(ethylene oxide) as sacrificial additive, followed by loading with norfloxacin and coating with an antifungal agent via dynamic imine bonds. Dynamic vapor sorption experiment proved intra-fiber mesopores around 2.7 nm, and UV-vis, FTIR, and NMR spectroscopy confirmed the norfloxacin embedding and the imination reaction. SEM, AFM and POM techniques displayed semicrystalline nanofibers with average diameter around 170 nm entangled into a non-woven mat. Their mesoporous nature favored a rapid adsorption of fluids up to 17 g/g, and a biodegradation rate fitting the wound healing rate, i.e. up to 30 % mass loss in media of pH characteristic to wound exudate and total degradation in that characteristic to normal dermis. The composite fibers released the NFX and 2FPBA in a controlled manner, and showed antimicrobial activity against gram positive, gram negative and fungal strains. They had no cytotoxic effect on normal human dermal fibroblasts, and showed biocompatibility on experimental rats. The investigation of wound healing ability on second/third-degree burn model in rats revealed wound closure and total restoration of the fully functional dermis and epidermis.

Quaternized chitosan (nano)fibers: A journey from preparation to high performance applications.

The industrial production of chitosan, initiated over 50 years ago, has transformed its application across diverse industries, agriculture, and medicine. To enhance its properties, numerous chitosan derivatives have been synthesized. The quaternization of chitosan has proven beneficial, as it not only enhances its properties but also imparts water solubility, expanding its potential for a wider range of applications. Specifically, the utilization of quaternized chitosan-based nanofibers has leveraged the synergistic benefits of quaternized chitosan (including hydrophilicity, bioadhesiveness, antimicrobial, antioxidant, hemostatic, and antiviral activities, as well as ionic conductivity) in combination with the distinctive characteristics of nanofibers (such as a high aspect ratio and 3D architecture). This combination has permitted numerous possibilities, spanning from wound dressings, air and water filters, drug delivery scaffolds, antimicrobial textiles, to energy storage systems and alkaline fuel cells. In this comprehensive review, we examine the preparation methods, properties, and applications of various composite fibers containing quaternized chitosan. The advantages and disadvantages of each method and composition are meticulously summarized, while relevant diagrams and figures illustrate the key findings.

Antitumor Activity of PEGylated and TEGylated Phenothiazine Derivatives: Structure-Activity Relationship.

The paper aims to investigate the antitumor activity of a series of phenothiazine derivatives in order to establish a structure-antitumor activity relationship. To this end, PEGylated and TEGylated phenothiazine have been functionalized with formyl units and further with sulfonamide units via dynamic imine bonds. Their antitumor activity was monitored in vitro against seven human tumors cell lines and a mouse one compared to a human normal cell line by MTS assay. In order to find the potential influence of different building blocks on antitumor activity, the antioxidant activity, the ability to inhibit farnesyltransferase and the capacity to bind amino acids relevant for tumor cell growth were investigated as well. It was established that different building blocks conferred different functionalities, inducing specific antitumor activity against the tumor cells.

Outstanding Sorption of Copper (II) Ions on Porous Phenothiazine-Imine-Chitosan Materials.

The aim of this work was to investigate the ability of a solid-state material, prepared by crosslinking chitosan with a phenothiazine-based aldehyde, to remove copper (II) ions from aqueous solutions, in a fast and selective manner. The metal uptake experiments, including the retention, sensibility, and selectivity against eight different metal ions, were realized via batch adsorption studies. The capacity of the material to retain copper (II) ions was investigated by spectrophotometric measurements, using poly(ethyleneimine) complexation agent, which allowed detection in a concentration range of 5-500 µM. The forces driving the copper sorption were monitored using various methods, such as FTIR spectroscopy, X-ray diffraction, SEM-EDAX technique, and optical polarized microscopy, and the adsorption kinetics were assessed by fitting the in vitro sorption data on different mathematical models. The phenothiazine-imine-chitosan material proved high ability to recover copper from aqueous media, reaching a maximum retention capacity of 4.394 g Cu (II)/g adsorbent when using a 0.5 M copper solution, which is an outstanding value compared to other chitosan-based materials reported in the literature to this date. It was concluded that the high ability of the studied xerogel to retain Cu (II) ions was the result of both physio- and chemo-sorption processes. This particular behavior was favored on one hand by the porous nature of the material and on the other hand by the presence of amine, hydroxyl, imine, and amide groups with the role of copper ligands.

Quaternized chitosan/chitosan nanofibrous mats: An approach toward bioactive materials for tissue engineering and regenerative medicine.

Chitosan based nanofibers are emerging biomaterials with a plethora of applications, especially in medicine and healthcare. Herein, binary quaternized chitosan/chitosan fibers are reported for the first time. Their preparation strategy consisted in the electrospinning of ternary chitosan/quaternized chitosan/poly(ethylene oxide) solutions followed by the selective removal of poly(ethylene oxide). Their morphology and performances were systematically investigated and discussed in detail. It was found that the fibers had reversible water vapor adsorption/desorption and showed swelling degrees similar to commercial wound dressings. They presented good mechanical properties and the content of quaternized chitosan modulated their bioadhesion, mucoadhesion and biodegradation rate and conferred them strong antimicrobial activity. Tests on normal human fibroblasts confirmed their safely use in contact with tissues and the biocompatibility investigation on rats showed no harmful effect when subcutaneous implanted. All these proved the binary quaternized chitosan/chitosan fibers as bioactive materials suitable for tissue regeneration, wound healing and drug delivery systems.

Microporous Polymelamine Framework Functionalized with Re(I) Tricarbonyl Complexes for CO2 Absorption and Reduction.

A mixture of polymeric complexes based on the reaction between Re(CO)5Cl and the porous polymeric network coming from the coupling of melamine and benzene-1,3,5-tricarboxaldehyde was obtained and characterized by FTIR, NMR, SEM, XPS, ICP, XRD, and cyclic voltammetry (CV). The formed rhenium-based porous hybrid material reveals a noticeable capability of CO2 absorption. The gas absorption amount measured at 295 K was close to 44 cm3/g at 1 atm. An interesting catalytic activity for CO2 reduction reaction (CO2RR) is observed, resulting in a turn over-number (TON) close to 6.3 under 80 min of test at -1.8 V vs. Ag/AgCl in a TBAPF6 0.1 M ACN solution. A possible use as filler in membranes or columns can be envisaged.

TEGylated Phenothiazine-Imine-Chitosan Materials as a Promising Framework for Mercury Recovery.

This paper reports new solid materials based on TEGylated phenothiazine and chitosan, with a high capacity to recover mercury ions from aqueous solutions. They were prepared by hydrogelation of chitosan with a formyl derivative of TEGylated phenothiazine, followed by lyophilization. Their structural and supramolecular characterization was carried out by 1H-NMR and FTIR spectroscopy, as well as X-ray diffraction and polarized light microscopy. Their morphology was investigated by scanning electron microscopy and their photophysical behaviour was examined by UV/Vis and emission spectroscopy. Swelling evaluation in different aqueous media indicated the key role played by the supramolecular organization for their hydrolytic stability. Mercury recovery experiments and the analysis of the resulting materials by X-ray diffraction and FTIR spectroscopy showed a high ability of the studied materials to bind mercury ions by coordination with the sulfur atom of phenothiazine, imine linkage, and amine units of chitosan.

Biocompatible drug delivery systems able to co-deliver antifungal and antiviral agents.

The present study reports the synthesis and characterization of 12 drug delivery systems (DDS) for the co-delivery of antifungal and antiviral agents. The systems were obtained by an in situ hydrogelation method of 6 chitosan oligomers with values of the polymerization degree between 14 and 51, with 2-formylphenylboronic acid, in the presence of tenofovir. The structural characterization by NMR and FTIR spectroscopy demonstrated the formation of imine linkages, while WXRD revealed the 3D layered architecture of the systems. SEM and POM images demonstrated the uniform distribution of tenofovir into the matrix, while the Zeta potential measurements revealed the strong interactions which develop between system components. The obtained DDSs presented biodegradability, hemocompatibility and in vivo biocompatibility, which along with their ability to release both the drug and the antifungal aldehyde make them promising materials for the treatment of HIV infection and its associated co-infections' symptoms.

Erythromycin Formulations-A Journey to Advanced Drug Delivery.

Erythromycin (ERY) is a macrolide compound with a broad antimicrobial spectrum which is currently being used to treat a large number of bacterial infections affecting the skin, respiratory tract, intestines, bones and other systems, proving great value from a clinical point of view. It became popular immediately after its discovery in 1952, due to its therapeutic effect against pathogens resistant to other drugs. Despite this major advantage, ERY exhibits several drawbacks, raising serious clinical challenges. Among them, the very low solubility in water and instability under acidic conditions cause a limited efficacy and bioavailability. Apart from this, higher doses promote drug resistance and undesirable effects. In order to overcome these disadvantages, during the past decades, a large variety of ERY formulations, including nanoparticles, have emerged. Despite the interest in ERY-(nano)formulations, a review on them is lacking. Therefore, this work was aimed at reviewing all efforts made to encapsulate ERY in formulations of various chemical compositions, sizes and morphologies. In addition, their preparation/synthesis, physico-chemical properties and performances were carefully analysed. Limitations of these studies, particularly the quantification of ERY, are discussed as well.

Self-Healing Chitosan Hydrogels: Preparation and Rheological Characterization.

The paper aims at the preparation of chitosan self-healing hydrogels, designed as carriers for local drug delivery by parenteral administration. To this aim, 30 hydrogels were prepared using chitosan and pyridoxal 5-phosphate (P5P), the active form of vitamin B6 as precursors, by varying the ratio of glucosamine units and aldehyde on the one hand and the water content on the other hand. The driving forces of hydrogelation were investigated by nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction, and polarized light microscopy (POM) measurements. NMR technique was also used to investigate the stability of hydrogels over time, and their morphological particularities were assessed by scanning electron microscopy (SEM). Degradability of the hydrogels was studied in media of four different pH, and preliminary self-healing ability was visually established by injection through a syringe needle. In-depth rheological investigation was conducted in order to monitor the storage and loss moduli, linear viscoelastic regime, and structural recovery capacity. It was concluded that chitosan crosslinking with pyridoxal 5-phosphate is a suitable route to reach self-healing hydrogels with a good balance of mechanical properties/structural recovery, good stability over time, and degradability controlled by pH.

Pegylation of phenothiazine - A synthetic route towards potent anticancer drugs.

Introduction: Cancer is a big challenge of the 21 century, whose defeat requires efficient antitumor drugs. Objectives: The paper aims to investigate the synergistic effect of two structural building blocks, phenothiazine and poly(ethylene glycol), towards efficient antitumor drugs. Methods: Two PEGylated phenothiazine derivatives were synthetized by attaching poly(ethylene glycol) of 550 Da to the nitrogen atom of phenothiazine by ether or ester linkage. Their antitumor activity has been investigated on five human tumour lines and a mouse tumor line as well, by determination of IC50. The in vivo toxicity was determined by measuring the LD50 in BALB/c mice by the sequential method and the in vivo antitumor potential was measured by the tumours growth test. The antitumor mechanism was investigated by complexation studies of zinc and magnesium ions characteristic to the farnesyltransferase enzyme, by studies of self-aggregation in the cells proximity and by investigation of the antitumor properties of the acid species resulted by enzymatic cleavage of the PEGylated derivatives. Results: The two compounds showed antitumor activity, with IC50 against mouse colon carcinoma cell line comparable with that of the traditional antitumor drugs 5-Fluorouracil and doxorubicin. The phenothiazine PEGylation resulted in a significant toxicity diminishing, the LD50 in BALB/c mice increasing from 952.38 up to 1450 mg/kg, in phenothiazine equivalents. Both compounds inflicted a 92% inhibition of the tumour growth for doses much smaller than LD50. The investigation of the possible tumour inhibition mechanism suggested the nanoaggregate formation and the cleavage of ester bonds as key factors for the inhibition of cancer cell proliferation and biocompatibility improvement. Conclusion: Phenothiazine and PEG building blocks have a synergetic effect working for both tumour growth inhibition and biocompatibility improvement. All these findings recommend the PEGylated phenothiazine derivatives as a valuable workbench for a next generation of antitumor drugs.

Liposomal-Based Formulations: A Path from Basic Research to Temozolomide Delivery Inside Glioblastoma Tissue.

Glioblastoma (GBM) is a lethal brain cancer with a very difficult therapeutic approach and ultimately frustrating results. Currently, therapeutic success is mainly limited by the high degree of genetic and phenotypic heterogeneity, the blood brain barrier (BBB), as well as increased drug resistance. Temozolomide (TMZ), a monofunctional alkylating agent, is the first line chemotherapeutic drug for GBM treatment. Yet, the therapeutic efficacy of TMZ suffers from its inability to cross the BBB and very short half-life (~2 h), which requires high doses of this drug for a proper therapeutic effect. Encapsulation in a (nano)carrier is a promising strategy to effectively improve the therapeutic effect of TMZ against GBM. Although research on liposomes as carriers for therapeutic agents is still at an early stage, their integration in GBM treatment has a great potential to advance understanding and treating this disease. In this review, we provide a critical discussion on the preparation methods and physico-chemical properties of liposomes, with a particular emphasis on TMZ-liposomal formulations targeting GBM developed within the last decade. Furthermore, an overview on liposome-based formulations applied to translational oncology and clinical trials formulations in GBM treatment is provided. We emphasize that despite many years of intense research, more careful investigations are still needed to solve the main issues related to the manufacture of reproducible liposomal TMZ formulations for guaranteed translation to the market.

Biocompatible Chitosan-Based Hydrogels for Bioabsorbable Wound Dressings.

Supramolecular hydrogels based on chitosan and monoaldehydes are biomaterials with high potential for a multitude of bioapplications. This is due to the proper choice of the monoaldehyde that can tune the hydrogel properties for specific practices. In this conceptual framework, the present paper deals with the investigation of a hydrogel as bioabsorbable wound dressing. To this aim, chitosan was cross-linked with 2-formylphenylboronic acid to yield a hydrogel with antimicrobial activity. FTIR, NMR, and POM procedures have characterized the hydrogel from a structural and supramolecular point of view. At the same time, its biocompatibility and antimicrobial properties were also determined in vitro. Furthermore, in order to assess the bioabsorbable character, its biodegradation was investigated in vitro in the presence of lysosome in media of different pH, mimicking the wound exudate at different stages of healing. The biodegradation was monitored by gravimetrical measurements, SEM microscopy and fractal analyses of the images. The fractal dimension values and the lacunarity of SEM pictures were accurately calculated. All these successful investigations led to the conclusion that the tested materials are at the expected high standards.

Chitosan crosslinking with a vanillin isomer toward self-healing hydrogels with antifungal activity.

The purpose of the study was to develop new antimicrobial hydrogels from natural resources that may promote wound healing and prevent bacterial skin infection. The new hydrogels were synthesized by crosslinking chitosan with a vanillin isomer, 5-methoxysalicylaldehyde, by a friendly and easy method. To characterize these hydrogels, their structural and morphological properties were explored by FTIR, 1H NMR, SEM, POM, and TGA. In view of the targeted application, swelling behavior, biodegradability, antimicrobial activity and biocompatibility were investigated in vitro. Structural and morphological studies confirmed the formation of new hydrogels via the imination reaction concomitant with the supramolecular organization. The hydrogels were highly porous with the average pore diameter around 80 µm, and a swelling rate controlled by the crosslinking density and medium pH. The hydrogels showed a progressive weight loss in the presence of lysozyme up to 35%, during 21 days of testing. They proved non-cytotoxic effect on Normal Human Dermal Fibroblasts using MTS test and powerful antifungal activity against Candida Albicans, as determined by disk diffusion assay. All these properties indicate the new hydrogels as a promising option for the treatment of various skin lesions.

Imination of Microporous Chitosan Fibers-A Route to Biomaterials with "On Demand" Antimicrobial Activity and Biodegradation for Wound Dressings.

Microporous chitosan nanofibers functionalized with different amounts of an antimicrobial agent via imine linkage were prepared by a three-step procedure including the electrospinning of a chitosan/PEO blend, PEO removal and acid condensation reaction in a heterogeneous system with 2-formylphenylboronic acid. The fibers' characterization was undertaken keeping in mind their application to wound healing. Thus, by FTIR and 1H-NMR spectroscopy, it was confirmed the successful imination of the fibers and the conversion degree of the amine groups of chitosan into imine units. The fiber morphology in terms of fiber diameter, crystallinity, inter- and intra-fiber porosity and strength of intermolecular forces was investigated using scanning electron microscopy, polarized light microscopy, water vapor sorption and thermogravimetric analysis. The swelling ability was estimated in water and phosphate buffer by calculating the mass equilibrium swelling. The fiber biodegradation was explored in five media of different pH, corresponding to different stages of wound healing and the antimicrobial activity against the opportunistic pathogens inflicting wound infection was investigated according to standard tests. The biocompatibility and bioadhesivity were studied on normal human dermal fibroblast cells by direct contact procedure. The dynamic character of the imine linkage of the functionalized fibers was monitored by UV-vis spectroscopy. The results showed that the functionalization of the chitosan microporous nanofibers with antimicrobial agents via imine linkage is a great route towards bio-absorbable wound dressings with "on demand" antimicrobial properties and biodegradation rate matching the healing stages.

Fluorescent chitosan-BODIPY macromolecular chemosensors for detection and removal of Hg2+ and Fe3+ ions.

The detection of heavy metals, such as Hg2+ and Fe3+, is of great significance. In this work, fluorescent small-molecule BODIPY (BY-3) bearing CC group was synthesized firstly. And then, the chitosan-based polymer sensor CY-1 was synthesized through the spontaneous NH2/C≡C click reaction. The synthesized CY-1 can effectively bind and recognize Hg2+/Hg+ by the -C=N groups formed in the click reaction. Moreover, the macromolecular sensors CS-1 and CS-2 were synthesized by incorporating another recognition sites to CY-1. These synthesized macromolecular sensors can not only recognize Hg2+/Hg+, but also effectively recognize Fe3+/Fe2+. All of them exhibited significant quenching effect, visible to the naked eye under UV irradiation. The detection limit of CY-1 for Hg2+ was 1.51 × 10-6 mol/L, and the detection limit of CS-2 for Fe3+ was 2.30 × 10-6 mol/L. The BODIPY-chitosan sensors synthesized in this work have the functions of removing heavy metal ions besides the identifying ability. The maximum adsorption capacity of 1 g chitosan to Hg2+ was 108 mg as the best one. This article provides a new method to prepare macromolecular sensors for the detection and removal of heavy metal ions. As a useful natural polymer, chitosan's application scope was enlarged.