Release Of Bovine Serum Albumin Research Articles

Triggered protein release from calcium alginate/chitosan gastro-resistant capsules

This work aimed to develop gastro-resistant capsules, based on alginate (Alg), as an oral delivery system for enhanced bioavailability of proteins. Bovine Serum Albumin (BSA), selected as a model protein, has been encapsulated inside calcium alginate capsules, which have been coated with a thin layer of chitosan (CHT) to improve its robustness and delivery profile. BSA and sodium alginate aqueous mixtures were characterized by optical microscopy and dynamic light scattering, suggesting that BSA and sodium alginate form water-in-water coacervate droplets by ionic complexation. Encapsulation of BSA-Alg colloidal dispersions was performed by introducing the BSA-Alg mixtures in CaCl2 solutions. Thus, calcium alginate capsules were formed, which were subsequently coated with CHT. Encapsulation of BSA achieved ≈99 % efficiency, the water content of capsules was ≈95 wt % and the amount of chitosan coating in wet capsules was <0.032 wt %. The stability of the capsules and the release of BSA were studied by several in vitro techniques, in simulated gastric and intestinal fluids. The addition of CHT as a coating material greatly improved the performance of the capsules, compared to that of capsules without CHT. The results of calcium alginate-chitosan capsules showed that they are stable in water at neutral pH and remain almost intact in simulated gastric fluid, but rapidly disintegrate when further treated with simulated intestinal fluid. These results demonstrated that calcium alginate-chitosan capsules can protect proteins from harsh gastric conditions and release their content at intestinal pH, demonstrating their potential application in the field of oral protein delivery.

PH-Responsive Cellulose/Silk/Fe3O4 Hydrogel Microbeads Designed for Biomedical Applications.

In this study, cellulose/Fe3O4 hydrogel microbeads were prepared through the sol-gel transition of a solvent-in-oil emulsion using various cellulose-dissolving solvents and soybean oil without surfactants. Particularly, 40% tetrabutylammonium hydroxide (TBAH) and 40% tetrabutylphosphonium hydroxide (TBPH) dissolved cellulose at room temperature and effectively dispersed Fe3O4, forming cellulose/Fe3O4 microbeads with an average diameter of ~15 µm. Additionally, these solvents co-dissolved cellulose and silk, allowing for the manufacture of cellulose/silk/Fe3O4 hydrogel microbeads with altered surface characteristics. Owing to the negatively charged surface characteristics, the adsorption capacity of the cellulose/silk/Fe3O4 microbeads for the cationic dye crystal violet was >10 times higher than that of the cellulose/Fe3O4 microbeads. When prepared with TBAH, the initial adsorption rate of bovine serum albumin (BSA) on the cellulose/silk/Fe3O4 microbeads was 18.1 times higher than that on the cellulose/Fe3O4 microbeads. When preparing TBPH, the equilibrium adsorption capacity of the cellulose/silk/Fe3O4 microbeads for BSA (1.6 g/g) was 8.5 times higher than that of the cellulose/Fe3O4 microbeads. The pH-dependent BSA release from the cellulose/silk/Fe3O4 microbeads prepared with TBPH revealed 6.1-fold slower initial desorption rates and 5.2-fold lower desorption amounts at pH 2.2 than those at pH 7.4. Cytotoxicity tests on the cellulose and cellulose/silk composites regenerated with TBAH and TBPH yielded nontoxic results. Therefore, cellulose/silk/Fe3O4 microbeads are considered suitable pH-responsive supports for orally administered protein pharmaceuticals.

The role of BSA protein on the skin regeneration ability of electrospun PCL/HA scaffolds, kinetics of release and essential in vitro tests

This study focused on the skin regeneration ability of hybrid polycaprolactone (PCL) scaffolds incorporated by hydroxyapatite (HA) nanoparticles (NPs) and bovine serum albumin (BSA). The scaffolds were synthesized by the electrospinning method, and their microstructure, molecular groups, wettability, swelling, porosity, and mechanical properties were characterized. The results showed that the HA NPs and BSA protein could be incorporated within the PCL fibers without a significant effect on the main molecular groups of the PCL structure. Incorporation of both the HA NPs and BSA protein could reduce the superhydrophobicity of the PCL scaffold to the required amount for skin regeneration. It was also found that the incorporation of mentioned elements increased the water uptake by PCL fibrous scaffolds, and decreased the mechanical properties. The kinetics of BSA release from the hybrid scaffolds was also investigated. The Higuchi model was the best-fitting model to explain the protein release from the electrospun products. The cytocompatibility of scaffolds was evaluated by the human umbilical vein endothelial cells (HUVEC), and the higher ability of the PCL/BSA scaffold for skin regeneration was confirmed.

Mussel inspired sequential protein delivery based on self-healing injectable nanocomposite hydrogel

Polysaccharide based self-healing and injectable hydrogels with reversible characteristics have widespread potential in protein drug delivery. However, it is a challenge to design the dynamic hydrogel for sequential release of protein drugs. Herein, we developed a novel mussel inspired sequential protein delivery dynamic polysaccharide hydrogel. The nanocomposite hydrogel can be fabricated through doping polydopamine nanoparticles (PDA NPs) into reversible covalent bond (imine bonds) crosslinked polymer networks of oxidized hyaluronic acid (OHA) and carboxymethyl chitosan (CEC), named PDA NPs@OHA-l-CEC. Besides multiple capabilities (i.e., injection, self-healing, and biodegradability), the nanocomposite hydrogel can achieve sustained and sequential protein delivery of vascular endothelial growth factor (VEGF) and bovine serum albumin (BSA). PDA NPs doped in hydrogel matrix serve dual roles, acting as secondary protein release structures and form dynamic non-covalent interactions (i.e., hydrogen bonds) with polysaccharides. Moreover, by adjusting the oxidation degree of OHA, the hydrogels with different crosslinking density could control overall protein release rate. Analysis of different release kinetic models revealed that Fickian diffusion drove rapid VEGF release, while the slower BSA release followed a Super Case II transport mechanism. The novel biocompatible system achieved sequential release of protein drugs has potentials in multi-stage synergistic drug deliver based on dynamic hydrogel.

Time-Delayed Anticancer Effect of an Extremely Low Frequency Alternating Magnetic Field and Multimodal Protein-Tannin-Mitoxantrone Carriers with Brillouin Microspectroscopy Visualization In Vitro.

The effect of an extremely low frequency alternating magnetic field (ELF AMF) at frequencies of 17, 48, and 95 Hz at 100 mT on free and internalized 4T1 breast cancer cell submicron magnetic mineral carriers with an anticancer drug, mitoxantrone, was shown. The alternating magnetic field (100 mT; 17, 48, 95 Hz; time of treatment-10.5 min with a 30 s delay) does not lead to the significant destruction of carrier shells and release of mitoxantrone or bovine serum albumin from them according to the data of spectrophotometry, or the heating of carriers in the process of exposure to magnetic fields. The most optimal set of factors that would lead to the suppression of proliferation and survival of cells with anticancer drug carriers on the third day (in comparison with the control and first day) is exposure to an alternating magnetic field of 100 mT in a pulsed mode with a frequency of 95 Hz. The presence of magnetic nanocarriers in cell lines was carried out by a direct label-free method, space-resolved Brillouin light scattering (BLS) spectrometry, which was realized for the first time. The analysis of the series of integrated BLS spectra showed an increase in the magnetic phase in cells with a growth in the number of particles per cell (from 10 to 100) after their internalization. The safety of magnetic carriers in the release of their constituent ions has been evaluated using atomic absorption spectrometry.

Effect of calcium phosphate/bovine serum albumin coated Al2O3-Ti biocomposites on osteoblast response.

The biological performance of aluminum oxide-titanium (Al2O3-Ti) composites requires special attention to achieve improved osteoblastic differentiation, and subsequent osseointegration/strong anchorage with the surrounding bone. Therefore, the aim of this study was to improve them by providing calcium phosphate (Ca-P)/bovine serum albumin (BSA) coating on their surfaces. Ca-P/BSA coatings were prepared on the surfaces of 75vol.%Ti composites (75Ti-BSA) and pure Ti (100Ti-BSA as a control). The surface characteristics, phase analysis, micro-hardness, BSA release profile and biological responses including cytotoxicity, cell viability, differentiation, mineralization, and cell adhesion were evaluated. The results showed that lower cytotoxicity% and higher mitochondrial activity or viability % were associated with the samples with Ca-P/BSA coatings (particularly 75Ti-BSA having 21.3% cytotoxicity, 111.4% and 288.6% viability at day 1 and 7, respectively). Furthermore, the Ca-P/BSA coating could highly enhance the differentiation of pre-osteoblast cells into osteoblasts in 75Ti-BSA group (ALP concentration of 4.8 ng/ml). However, its influence on cell differentiation in 100Ti-BSA group was negligible. Similar results were also obtained from mineralization assay. The results on cell adhesion revealed that the Ca-P/BSA coated samples differently interacted with MC3T3-E1 cells; enlarged flat cells on 75Ti-BSA vs more spindle-shaped cells on 100Ti-BSA. Ca-P/BSA coated Al2O3-Ti provided promising biological performance, superior to that of uncoated composites. Therefore, they have the potential to improve implant osseointegration.

Poly(N-isopropylacrylamide) based smart nanofibrous scaffolds for use as on-demand delivery systems for oral and dental tissue regeneration.

Stimuli-responsive domains capable of releasing loaded molecules, "on-demand," have garnered increasing attention due to their enhanced delivery, precision targeting, and decreased adverse effects. The development of an on-demand delivery system that can be easily triggered by dental clinicians might have major roles in dental and oral tissue engineering. A series of random graft poly(NIPAm-co-HEMA-Lactate) copolymers were synthesized using 95:5, 85:5, 60:40, and 40:60 ratios of thermosensitive NIPAm and HEMA-poly lactate respectively then electrospun to produce nanofibrous scaffolds loaded with bovine serum albumin (BSA). Cumulative BSA release was assessed at 25C and 37°C. To appraise the use of scaffolds as on-demand delivery systems, they were subjected to thermal changes in the form cooling and warming cycles during which BSA release was monitored. To confirm the triggered releasing ability of the synthesized scaffolds, the copolymer made with 60% NIPAm was selected, based on the results of the release tests, and loaded with bone morphogenetic protein-2 (BMP-2). The loaded scaffolds were placed with mesenchymal-like stem cells (iMSCs) derived from induced pluripotent stem cells (iPSCs), and subjected to temperature alterations. Then, the osteogenic differentiation of iMSCs, which might have resulted from the released protein, was evaluated after 10 days by analyzing runt-related transcription factor 2 (RUNX-2) osteogenic gene expression by the cells using real-time quantitative polymerase chain reaction (qRT-PCR). BSA release profiles showed a burst release at the beginning followed by a more linear pattern at 25°C, and a much slower release at 37°C. The release also decreased when the PNIPAm content decreased in the scaffolds. Thermal triggering led to a step-like release pattern in which the highest release was reported 30 min through the warming cycles. The iMSCs cultivated with scaffolds loaded with BMP-2 and exposed to temperature alteration showed significantly higher RUNX-2 gene expression than cells in the other experimental groups. The synthesized scaffolds are thermo-responsive and could be triggered to deliver biological biomolecules to be used in oral and dental tissue engineering. Thermal stimuli could be simulated by dental clinicians using simple means of cold therapy, for example, cold packs in intraoral accessible sites for specified times.

Unraveling mechanisms of protein encapsulation and release in coacervates via molecular dynamics and machine learning.

Coacervates play a pivotal role in protein-based drug delivery research, yet their drug encapsulation and release mechanisms remain poorly understood. Here, we utilized the Martini model to investigate bovine serum albumin (BSA) protein encapsulation and release within polylysine/polyglutamate (PLys/PGlu) coacervates. Our findings emphasize the importance of ingredient addition sequence in coacervate formation and encapsulation rates, attributed to preference contact between oppositely charged proteins and poly(amino acid)s. Notably, coacervates composed of β-sheet poly(amino acid)s demonstrate greater BSA encapsulation efficiency due to their reduced entropy and flexibility. Furthermore, we examined the pH responsiveness of coacervates, shedding light on the dissolution process driven by Coulomb forces. By leveraging machine learning algorithms to analyze simulation results, our research advances the understanding of coacervate-based drug delivery systems, with the ultimate goal of optimizing therapeutic outcomes.

Exfoliated clay-polymer nanocomposite as a promising delivery system for bovine serum albumin

ABSTRACT The pH-sensitive exfoliated clay-alginate composite was investigated for the controlled oral delivery of bovine serum albumin (BSA). The X-ray diffraction analysis and transmission electron microscopy analysis revealed the exfoliated structure of clay in the composite. The degree of exfoliation was increased with sonication and it was optimised that the maximum exfoliation of clay in the aqueous medium occurred for 150 min of sonication. The maximum adsorption of BSA in the clay-alginate was detected at a pH of 4.5. The in-vitro release study of BSA from the exfoliated clay and the clay-alginate composite was analysed at simulated gastric and simulated intestinal pH conditions which suggested that the adsorption and release of BSA could be controlled simply by adjusting the solution pH condition. In the in-vitro release study, it was noted that there was no release of BSA from the clay-alginate composite up to 30 min in gastric pH conditions. But, there was a slow release rate of BSA within 10 h from the clay-alginate composite in the intestinal pH condition. The release profile from the clay–alginate composite was more fitted to the Peppas kinetic model.

Guar-Based Injectable Hydrogel for Drug Delivery and In Vitro Bone Cell Growth.

Injectable hydrogels offer numerous advantages in various areas, which include tissue engineering and drug delivery because of their unique properties such as tunability, excellent carrier properties, and biocompatibility. These hydrogels can be administered with minimal invasiveness. In this study, we synthesized an injectable hydrogel by rehydrating lyophilized mixtures of guar adamantane (Guar-ADI) and poly-β-cyclodextrin (p-βCD) in a solution of phosphate-buffered saline (PBS) maintained at pH 7.4. The hydrogel was formed via host-guest interaction between modified guar (Guar-ADI), obtained by reacting guar gum with 1-adamantyl isocyanate (ADI) and p-βCD. Comprehensive characterization of all synthesized materials, including the hydrogel, was performed using nuclear magnetic resonance (NMR) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and rheology. The in vitro drug release study demonstrated the hydrogel's efficacy in controlled drug delivery, exemplified by the release of bovine serum albumin (BSA) and anastrozole, both of which followed first-order kinetics. Furthermore, the hydrogel displayed excellent biocompatibility and served as an ideal scaffold for promoting the growth of mouse osteoblastic MC3T3 cells as evidenced by the in vitro biocompatibility study.

Preparation and Properties of Sodium Alginate/Gelatin Hydrogel-Microspheres

Calcium alginate gel microspheres exhibiting pH response release characteristics were prepared, and utilizing Bovine serum albumin (BSA) to evaluate drug release performance. Hydrogels composed of gelatin (GL) and sodium alginate oxide (NALO), with varying ratios (7:3; 4:1; 3:2) were prepared by controlling their composition. Scanning electron microscopy (SEM) and a microplate reader (ELIASA) were used to characterize the microspheres’ microstructure and in vitro BSA release behavior, respectively. The results demonstrated that, under acidic conditions, the BSA release was slower than in neutral conditions. The hydrogels’ physical and chemical properties were analyzed using optical microscopy, Fourier transform infrared spectroscopy (FTIR), swelling experiments and mechanical testing of the hydrogels and microsphere loaded hydrogels. The results showed that a GL to NALO ratio of 7:3 increased the swelling degree, while a 4:1 ratio optimized the mechanical characteristics. The study concluded that the microsphere-loaded hydrogels exhibited significant potential for human drug delivery applications.

A Single Injection with Sustained-Release Microspheres and a Prime-Boost Injection of Bovine Serum Albumin Elicit the Same IgG Antibody Response in Mice

Although vaccination is still considered to be the cornerstone of public health care, the increase in vaccination coverage has stagnated for many diseases. Most of these vaccines require two or three doses to be administered across several months or years. Single-injection vaccine formulations are an effective method to overcome the logistical barrier to immunization that is posed by these multiple-injection schedules. Here, we developed subcutaneously (s.c.) injectable microspheres with a sustained release of the model antigen bovine serum albumin (BSA). The microspheres were composed of blends of two novel biodegradable multi-block copolymers consisting of amorphous, hydrophilic poly(ε-caprolactone)-poly(ethylene glycol)-poly(ε-caprolactone) (PCL-PEG-PCL) blocks and semi-crystalline poly(dioxanone) (PDO) blocks of different block sizes. In vitro studies demonstrated that the release of BSA could be tailored over a period of approximately four to nine weeks by changing the blend ratio of both polymers. Moreover, it was found that BSA remained structurally intact during release. Microspheres exhibiting sustained release of BSA for six weeks were selected for the in vivo study in mice. The induced BSA-specific IgG antibody titers increased up to four weeks after administration and were of the same magnitude as found in mice that received a priming and a booster dose of BSA in phosphate-buffered saline (PBS). Determination of the BSA concentration in plasma showed that in vivo release probably took place up to at least four weeks, although plasma concentrations peaked already one week after administration. The sustained-release microspheres might be a viable alternative to the conventional prime-boost immunization schedule, but a clinically relevant antigen should be incorporated to assess the full potential of these microspheres in practice.

In vitro Characterization of Polyethyleneimine-Oleic Acid Cationic Micelle as a Novel Protein Carrier.

Nanotechnology has introduced valuable carriers for vaccine delivery. The success of vaccination depends on many factors, such as the intact and safe presentation of vaccine candidates to immune cells. We have conjugated branched PEI-2k and oleic acid (OL) as the building block of the cationic micelle. We aimed to introduce a novel carrier for vaccine candidates. We conjugated polyethyleneimine and OL (POA) to synthesize the building blocks of cationic micelles. The critical micelle concentration (CMC), size and zeta potential of micelles, and their stability in 60 days were determined. Loading, encapsulation efficiency, and in vitro release study were assessed using bovine serum albumin (BSA) as a protein model. Furthermore, the cytotoxicity and hemocompatibility of developed nanosized micelles were evaluated to ascertain the biocompatibility of fabricated micelles. Cell uptake of cationic micelles in the macrophage cell line was also followed up. The conjugation of two polymer parts was confirmed by Fourier transform infrared spectroscopy and 1H nuclear magnetic resonance techniques. The CMC of the developed micelles was around 5.62 × 10-8 mg/ ml, whereas the loading and encapsulation efficiencies were 16.5% and 70%, respectively. The size and zeta potential of the cationic micelles were 96.53 ± 18.53 nm and 68.3 mV, respectively. The release of BSA from POA micelles after 8 and 72 hours was 8.5% and 82%, respectively. Finally, fluorescence microscopy showed that the prepared micelles were successfully and effectively taken up by RAW264.7 cells. These results may provide a cutting-edge vaccine delivery solution and open up a new avenue for future vaccine research.

Gelatin/alginate-based microspheres with sphere-in-capsule structure for spatiotemporal manipulative drug release in gastrointestinal tract

Microsphere with sphere-in-capsule structure is a multi-drugs delivery system to achieve the purpose of combination therapy. In this paper, we have prepared gelatin/alginate-based microspheres with sphere-in-capsule structure by a relatively fast, simple, and easily large-scale industrialized emulsification method for spatiotemporal manipulative drug release in gastrointestinal tract. Calcium alginate microspheres encapsulated with bovine serum albumin (BSA) were first prepared as inner microspheres, and then inner microspheres and ranitidine hydrochloride (RH) were co-encapsulated by gelatin microspheres to form double-layer microspheres with sphere-in-capsule structure. The size and distribution of microspheres can be easily controlled by emulsifying conditions. The microspheres with sphere-in-capsule structure displayed desirable encapsulation efficiency of BSA (61.52 %) and RH (56.07 %). The in vitro simulated drug release showed the spatiotemporal release feature of microspheres with sphere-in-capsule structure. In the specific simulated fluid, the release behavior and cumulative release of RH (sustainedly released 95 % in simulated gastric fluid) and BSA (rapidly released 73 % in simulated intestinal fluid) were different. The drug release mechanisms were analyzed to determine RH and BSA's release behavior. Overall, the microspheres with sphere-in-capsule structure have the potential application of spatiotemporal manipulative drug delivery in the gastrointestinal tract.

Albumin-loaded thermo/pH dual-responsive nanogels based on sodium alginate and poly (N-vinyl caprolactam).

During the past decades, many researchers have tried to encapsulate medicines in biopolymer nanogels as injectable medicines. In the present study, dual-responsive bovine serum albumin (BSA)-loaded nanogels prepared from sodium alginate grafted poly (N-vinyl caprolactam) (PNVCL) have been reported. First, PNVCL-g-sodium alginate (PNVCL-g-Alg) was synthesized through free radical polymerization, and then nanogels were obtained from ionic crosslinking of sodium alginate in the presence of BSA. FTIR spectra showed that PNVCL-g-Alg nanogels were successfully prepared. Turbidimetry and rheometry analyses demonstrated the cloud point temperature near the human body. Particle size was evaluated using FE-SEM and dynamic light scattering and it was found that the size of particles in dry and swollen state are about 30 and 280nm, respectively. The effect of temperature and pH on BSA release was evaluated. By comparing the drug release behavior, we found that the release of the protein at the temperature above the cloud point is faster than that at the temperature below the cloud point. The pH sensitivity of BSA-loaded PNVCL-g-Alg was evaluated at pH 5.5 and 7.4 and showed that the drug release was faster at acidic pH than at neutral pH.

Preparation and Characterization of PLG Microparticles by the Multiple Emulsion Method for the Sustained Release of Proteins.

Rapid release and diminished stability are two of the limitations associated with the growth factors that are essentially used in dental applications. These growth factors are employed to enhance the quality and quantity of tissue or bone matter during regeneration. Therefore, drug delivery devices and systems have been developed to address these limitations. In this study, bovine serum albumin (BSA), as a representative growth factor, was successfully sustained by encapsulation with the medium-absorbable copolymer, poly(L-lactide-co-glycolide) (PLG) 70:30% mol, via the multiple emulsion method. Different PLG, PVA, and BSA concentrations were used to investigate their effects on the BSA encapsulation efficiency. The suitable ratios leading to a better characterization of microparticles and a higher encapsulation efficiency in producing encapsulated PLG microparticles were 8% (w/v) of PLG, 0.25% (w/v) of PVA, and 8% (w/v) of BSA. Furthermore, an in vitro release study revealed a bursting release of BSA from the encapsulated PLG microsphere in the early phase of development. Subsequently, a gradual release was observed over a period of eight weeks. Furthermore, to encapsulate LL-37, different proteins were used in conjunction with PLG under identical conditions with regard to the loading efficiency and morphology, thereby indicating high variations and poor reproducibility. In conclusion, the encapsulated PLG microparticles could effectively protect the protein during encapsulation and could facilitate sustainable protein release over a period of 60 days. Importantly, an optimal method must be employed in order to achieve a high degree of encapsulation efficiency for all of the protein or growth factors. Accordingly, the outcomes of this study will be useful in the manufacture of drug delivery devices that require medium-sustained release growth factors, particularly in dental treatments.

Dually responsive biodegradable drug releasing 3D printed structures

AbstractThe aim of this study was to develop temperature and pH‐responsive biodegradable 3D printed drug‐releasing constructs using the Digital Light Processing (DLP) printing technique. The printable molecules, named “inks” throughout this study, comprised polyethylene oxide–polypropylene oxide–polyethylene oxide (PEO–PPO–PEO) triblocks and acrylic acid (AAc) that rendered them reverse thermo‐responsive (RTR) and pH‐sensitive, respectively. Additionally, the inks' biodegradability was imparted to them by incorporating lactide (LA) and ε‐caprolactone (CL) units into their composition, while their printability was achieved by end‐capping them with 2‐Isocyanatoethyl methacrylate (IEMA). By fine tuning the LA/CL composition, a broad range of inks displaying different water absorption behavior, biodegradation as well as bovine serum albumin (BSA) release profiles at different conditions. All the hydrogels showed a fast and reversible swelling–deswelling response, as they fluctuated between pH 2.0 and pH 7.4 or between 10 and 37°C. The resultant hydrogels showed good water uptake capacity (6340%) at pH 7.4 and 10°C temperature. BSA release from the loaded 3D printed hydrogel showed a maximum (80.5%) at pH 7.4 and 10°C in comparison to pH 7.4 and 37°C (65%). The proposed dual responsive 3D printed biodegradable hydrogels objects will shed light as a drug carrier.

A TGF-α and TGF-β1 Poloxamer-based micelle/hydrogel composite: A promising novel candidate for the treatment of anosmia.

Anosmia is the inability to smell or loss of the sense of smell. It can reduce your ability to detect the smell of smoke, gas leaks, or spoiled food, as well as hinder the quality of life related to social interactions and feelings of well-being. In the current study, a drug delivery composite was designed to cure anosmia and its efficiency in delivering transforming growth factor alpha (TGF-α) and transforming growth factor beta 1 (TGF-β1) to the nasal cavity was evaluated. Bovine serum albumin (BSA) was used as a model protein for encapsulation into Poloxamers 407 micelles. For the optimization of the BSA-micelle formulation, a two-parameter five-level central composite design (CCD) was applied. The BSA-micelle was optimized with a particle size of 41 nm, drug loading of 8%, and encapsulation efficiency of 74%. Further, the BSA-micelle was characterized by FESEM, TEM, and FTIR. The analysis of release profile suggested high-paced free BSA release compared to the gradual and prolonged release of BSA-micelle/hydrogel and BSA-micelles. The cytotoxicity assay demonstrated the safety of TGF-α and TGF-β1-micelles/hydrogel. Moreover, it was observed that TGF-α and TGF-β1 within the hydrogels promote cellular viability and human olfactory ectomesenchymal stem cell OE-MSCs proliferation. In conclusion, According to the results of our study, the TGF-α and TGF-β1-micelle/hydrogel-based delivery system provides a suitable alternative for anosmia treatment.

Formulation and Characterization of Alginate Coated Chitosan Nanoparticles as Therapeutic Protein for Oral Delivery System

Nanoparticles have been promptly studied and developed for oral protein delivery. Selection of excipients and formulation method depends on the physicochemical characteristics of the protein carried. Therefore, this study aims to design a formulation and characterize the alginate coated chitosan nanoparticles which carried different protein. Alginate coated chitosan nanoparticles were formed using an ionic gelation method. Optimization was done by varying the parameter such as crosslinker concentration, agitation method, rate, and time. The results show that chitosan nanoparticles formed by sonication using sodium tripolyphosphate (STPP): chitosan (1:0.8) was the best method to form nanoparticles. The particle size and polydispersity index (PDI) of bovine serum albumin (BSA) and lysozyme nanoparticles were 812.2 nm and 0.412 for BSA while 793.3 nm and 0.438 for lysozyme. Encapsulation efficiency (EE) of BSA is 52 % and lysozyme is 68 %. In vitro tests on acidic/gastric conditions showed that lysozyme (±32 %) was released faster than BSA (±10 %) during the 24 h incubation. Under neutral/terminal intestine conditions, percentage of BSA (±17 %) release is slightly higher than lysozyme (±13 %) for 8 h incubation period. It was concluded that the formulation of alginate coated chitosan nanoparticles in this study appears to be more effective for BSA delivery than lysozyme.&#x0D; HIGHLIGHTS&#x0D; &#x0D; Nanoparticle vesicular system is an effective approach for protein drug delivery system&#x0D; Factors affecting the formation of chitosan nanoparticle are crosslinker concentration, agitation method, rate, and time&#x0D; Alginate coated chitosan nanoparticle was more effective for oral delivery of BSA compared to lysozyme&#x0D; &#x0D; GRAPHICAL ABSTRACT

Manufacturing of self-standing multi-layered 3D-bioprinted alginate-hyaluronate constructs by controlling the cross-linking mechanisms for tissue engineering applications

Three-dimensional (3D) bioprinting of self-supporting stable tissue and organ structure is critically important in extrusion-based bioprinting system, especially for tissue engineering and regenerative medicine applications. However, the development of self-standing bioinks with desired crosslinking density, biocompatibility, tunable mechanical strength and other properties like self-healing, in situ gelation, drug or protein incorporation is still a challenge. In this study, we report a hydrogel bioink prepared from alginate (Alg) and hyaluronic acid (HA) crosslinked through multiple crosslinking mechanisms, i.e. acyl-hydrazone, hydrazide interactions and calcium ions. These Alg-HA gels were highly dynamic and shear-thinning with exceptional biocompatibility and tunable mechanical properties. The increased dynamic nature of the gels is mainly chemically attributed to the presence of acyl-hydrazone bonds formed between the amine groups of the acyl-hydrazide of alginate and the monoaldehyde of the HA. Among the different combinations of Alg-HA gel compositions prepared, the A5H5 (Alginate-acyl-hydrazide:HA-monoaldehyde, ratio 50:50) gel showed a gelation time of ∼60 s, viscosity of ∼400 Pa s (at zero shear rate), high stability in various pH solutions and increased degradation time (>50 days) than the other samples. The A5H5 gels showed high printability with increased post-printing stability as observed from the 3D printed structures (e.g. hollow tube (∼100 layers), porous cube (∼50 layers), star, heart-in, meniscus and lattice). The scanning electron microscopy analysis of the 3D constructs and hydrogels showed the interconnected pores (∼181 µm) and crosslinked networks. Further, the gels showed sustained release of 5-amino salicylic acid and bovine serum albumin. Also, the mechanical properties were tuned by secondary crosslinking via different calcium concentrations. In vitro assays confirmed the cytocompatibility of these gels, where the 3D bioprinted lattice and tubular (∼70 layers) constructs demonstrated high cell viability under fluorescence analysis. In in vivo studies, Alg-HA gel showed high biocompatibility (>90%) and increased angiogenesis (threefolds) and reduced macrophage infiltration (twofold decrease), demonstrating the promising potential of these hydrogels in 3D bioprinting applications for tissue engineering and regenerative medicine with tunable properties.