From Basic Principles of Protein-Polysaccharide Association to the Rational Design of Thermally Sensitive Materials.

Biology resolves design requirements toward functional materials by creating nanostructured composites, where individual components are combined to maximize the macroscale material performance. A major challenge in utilizing such design principles is the trade-off between the preservation of individual component properties and emerging composite functionalities. Here, polysaccharide pectin and silk fibroin were investigated in their composite form with pectin as a thermal-responsive ion conductor and fibroin with exceptional mechanical strength. We show that segregative phase separation occurs upon mixing, and within a limited compositional range, domains ∼50 nm in size are formed and distributed homogeneously so that decent matrix collective properties are established. The composite is characterized by slight conformational changes in the silk domains, sequestering the hydrogen-bonded ß-sheets as well as the emergence of randomized pectin orientations. However, most dominant in the composite's properties is the introduction of dense domain interfaces, leading to increased hydration, surface hydrophilicity, and increased strain of the composite material. Using controlled surface charging in X-ray photoelectron spectroscopy, we further demonstrate Ca ions (Ca2+) diffusion in the pectin domains, with which the fingerprints of interactions at domain interfaces are revealed. Both the thermal response and the electrical conductance were found to be strongly dependent on the degree of composite hydration. Our results provide a fundamental understanding of the role of interfacial interactions and their potential applications in the design of material properties, polysaccharide-protein composites in particular.

Design and fabrication of a magnetic nanobiocomposite based on flaxseed mucilage hydrogel and silk fibroin for biomedical and in-vitro hyperthermia applications.

In this research work, a magnetic nanobiocomposite is designed and presented based on the extraction of flaxseed mucilage hydrogel, silk fibroin (SF), and Fe3O4 magnetic nanoparticles (Fe3O4 MNPs). The physiochemical features of magnetic flaxseed mucilage hydrogel/SF nanobiocomposite are evaluated by FT-IR, EDX, FE-SEM, TEM, XRD, VSM, and TG technical analyses. In addition to chemical characterization, given its natural-based composition, the in-vitro cytotoxicity and hemolysis assays are studied and the results are considerable. Following the use of highest concentration of magnetic flaxseed mucilage hydrogel/SF nanobiocomposite (1.75 mg/mL) and the cell viability percentage of two different cell lines including normal HEK293T cells (95.73%, 96.19%) and breast cancer BT549 cells (87.32%, 86.9%) in 2 and 3 days, it can be inferred that this magnetic nanobiocomposite is biocompatible with HEK293T cells and can inhibit the growth of BT549 cell lines. Besides, observing less than 5% of hemolytic effect can confirm its hemocompatibility. Furthermore, the high specific absorption rate value (107.8 W/g) at 200 kHz is generated by a determined concentration of this nanobiocomposite (1 mg/mL). According to these biological assays, this magnetic responsive cytocompatible composite can be contemplated as a high-potent substrate for further biomedical applications like magnetic hyperthermia treatment and tissue engineering.

Self-assembled silk fibroin cross-linked with genipin supplements microbial carbonate precipitation in building material.

The process of microbially induced carbonate precipitation (MICP) is known to effectively improve engineering properties of building materials and so does silk fibroin (SF). Thus, in this study, an attempt was taken to see the improvement in sand, that is, basic building material coupled with MICP and SF. Urease producing Bacillus megaterium was utilized for MICP in Nutri-Calci medium. To improve the strength of SF itself in bacterial solution, it was cross-linked with genipin at the optimized concentration of 3.12 mg/mL. The Fourier transform infrared (FTIR) spectra confirmed the crosslinking of SF with genipin in bacterial solution. In order to understand how such cross-linking can improve engineering properties, sand moulds of 50 mm3 dimension were prepared that resulted in 35% and 55% more compressive strength than the one prepared with bacterial solution with SF and bacterial solution only, respectively with higher calcite content in former one. The FTIR, SEM, x-ray powder diffraction spectrometry and x-ray photoelectron spectroscopy analyses confirmed higher biomineral precipitation in bacterial solution coupled with genipin cross-linked SF. As the process of MICP is proven to replace cement partially from concrete without negatively influence mechanical properties, SF cross-linked with genipin can provide additional significance in developing low-carbon cement-based composites.

Magnetic xanthan gum-silk fibroin hydrogel: A nanocomposite for biological and hyperthermia applications.

A magnetic xanthan hydrogel/silk fibroin nanobiocomposite (XG hydrogel/SF/Fe3O4) was designed, fabricated, and characterized using analyzing methods such as FT-IR, EDX, FE-SEM, XRD, TGA, and VSM to evaluate the exact structure of product nanobiocomposite. The FE-SEM images reveal the presence of spherical shapes exhibiting a narrow size range and homogeneous distribution, measuring between 30 and 35 nm in diameter. The VSM analysis demonstrates the superparamagnetic properties of the XG hydrogel/SF/Fe3O4 nanobiocomposite, exhibiting a magnetic saturation of 54 emu/g at room temperature. The biological response of the nanobiocomposite scaffolds was assessed through cell viability and red blood cell hemolytic assays. MCF10A cells were exposed to a concentration of 1.75 mg/mL of the nanobiocomposite, and after 2 and 3 days, the cell viability was found to be 96.95 % and 97.02 %, respectively. The hemolytic effect was nearly 0 % even at higher concentrations (2 mg/mL). Furthermore, the magnetic nanobiocomposite showed excellent potential for hyperthermia applications, with a maximum specific absorption rate of 7 W/g for 1 mg/mL of the sample under a magnetic field in different frequencies (100, 200, 300, and 400 MHz) and 5 to 20 min time intervals.

Active colorimetric bilayer polycaprolactone-eucalyptus oil@silk fibroin-bayberry anthocyanins (PCL-EO@SF-BAs) membrane with directional water transport (DWT) for food packaging.

Currently, designing smart membranes with multifunctional effectiveness is crucial to food freshness monitoring and retention. Herein, an active colorimetric Janus bilayer membrane with directional water transport (DWT) performance is constructed by electrospinning, which comprises a hydrophilic layer of silk fibroin-bayberry anthocyanins (SF-BAs) and a hydrophobic layer of polycaprolactone-eucalyptus oil (PCL-EO). The entities of BAs and EO are well dispersed in the fiber matrix by hydrogen bonds and physical interactions, respectively. BAs endow the membrane colorimetric response and antioxidant activity, and EO contributes to the antibacterial activity while DWT performance is generated from the asymmetric wettability of the two layers. The bilayer membrane has an accumulative one-way transport index of 1077%, an overall moisture management capacity of 0.76 and a water evaporation rate of 0.48 g h-1. Moreover, the release of BAs and EO was predominantly controlled by Fickian diffusion. As a pH-sensing indicator, PCL-EO@SF-BAs is highly sensitive to external pH stimuli and the response is reversible. In addition to freshness monitoring, PCL-EO@SF-BAs can extend the shelf-life of pork beyond 100% at 4 °C. Also, it can extend the shelf life of shrimp by approximately 70% at 25 °C with the synergistic effect of antibacterial activity and the DWT performance.

A high molecular weight silk fibroin scaffold that resists degradation and promotes cell proliferation.

The regulation of the biodegradation rate of 3D-regenerated silk fibroin scaffolds and the avoidance of premature collapse are important concerns for their effective applications in tissue engineering. In this study, bromelain, which is specific to sericin, was used to remove sericin from silk, and high molecular weight silk fibroin was obtained after the fibroin fibers were dissolved. Afterwards, a 3D scaffold was prepared via freeze-drying. The Sodium dodecyl sulfate-polyacrylamide gel electrophoresis results showed that the average molecular weight of the regenerated silk fibroin prepared by using the bromelain-degumming method was approximately 142.2 kDa, which was significantly higher than that of the control groups prepared by using the urea- and Na2 CO3 -degumming methods. The results of enzyme degradation in vitro showed that the biodegradation rate and internal three-dimensional structure collapse of the bromelain-degumming fibroin scaffolds were significantly slower than those of the two control scaffolds. The proliferation activity of human umbilical vein vascular endothelial cells inoculated in bromelain-degumming fibroin scaffolds was significantly higher than that of the control scaffolds. This study provides a novel preparation method for 3D-regenerated silk fibroin scaffolds that can effectively resist biodegradation, continuously guide cell growth, have good biocompatibility, and have the potential to be used for the regeneration of various connective tissues.

Biomedical applications of chitosan/silk fibroin composites: A review.

Natural polymers have been used in the biomedical fields for decades, mainly derived from animals and plants with high similarities with biomacromolecules in the human body. As an alkaline polysaccharide, chitosan (CS) attracts much attention in tissue regeneration and drug delivery with favorable biocompatibility, biodegradation, and antibacterial activity. However, to overcome its mechanical properties and degradation behavior drawbacks, a robust fibrous protein-silk fibroin (SF) was introduced to prepare the CS/SF composites. Not only can CS be combined with SF via the amide and hydrogen bond formation, but also their functions are complementary and tunable with the blending ratio. To further improve the performances of CS/SF composites, natural (e.g., hyaluronic acid and collagen) and synthetic biopolymers (e.g., polyvinyl alcohol and hexanone) were incorporated. Also, the CS/SF composites acted as slow-release carriers for inorganic non-metals (e.g., hydroxyapatite and graphene) and metal particles (e.g., silver and magnesium), which could enhance cell functions, facilitate tissue healing, and inhibit bacterial growth. This review presents the state-of-the-art and future perspectives of different biomaterials combined with CS/SF composites as sponges, hydrogels, membranes, particles, and coatings. Emphasis is devoted to the biological potentialities of these hybrid systems, which look rather promising toward a multitude of applications.

Magnetic carboxymethyl cellulose-silk fibroin hydrogel: A ternary nanobiocomposite exhibiting excellent biological activity and in vitro hyperthermia of cancer therapy.

In this work, a magnetic nanobiocomposite scaffold based on carboxymethylcellulose (CMC) hydrogel, silk fibroin (SF), and magnetite nanoparticles was fabricated. The structural properties of this new magnetic nanobiocomposite were characterized by various analyses such as FT-IR, XRD, EDX, FE-SEM, TGA and VSM. According to the particle size histogram, most of the particles were between 55 and 77 nm and the value of saturation magnetization of this nanobiocomposite was reported 41.65 emu.g- 1. Hemolysis and MTT tests showed that the designed magnetic nanobiocomposite was compatible with the blood. In addition, the viability percentage of HEK293T normal cells did not change significantly, and the proliferation rate of BT549 cancer cells decreased in its vicinity. EC50 values for HEK293T normal cells after 48 h and 72 h were 3958 and 2566, respectively. Also, these values for BT549 cancer cells after 48 h and 72 h were 0.4545 and 0.9967, respectively. The efficiency of fabricated magnetic nanobiocomposite was appraised in a magnetic fluid hyperthermia manner. The specific absorption rate (SAR) of 69 W/g (for the 1 mg/mL sample at 200 kHz) was measured under the alternating magnetic field (AMF).

Injectable methacrylated gelatin/thiolated pectin hydrogels carrying melatonin/tideglusib-loaded core/shell PMMA/silk fibroin electrospun fibers for vital pulp regeneration.

Use of injectable hydrogels attract attention in the regeneration of dental pulp due to their ability to fill non-uniform voids such as pulp cavities. Here, gelatin methacrylate/thiolated pectin hydrogels (GelMA/PecTH) carrying electrospun core/shell fibers of melatonin (Mel)-polymethylmethacrylate (PMMA)/Tideglusib (Td)-silk fibroin (SF) were designed as an injectable hydrogel for vital pulp regeneration, through prolonged release of Td and Mel to induce proliferation and odontoblastic differentiation of dental pulp stem cells (DPSC). H NMR and FTIR confirmed methacrylation of Gel and thiolation of Pec. Addition of PMMA/SF increased degradation and water retention capacities of GelMA/PecTH. Rheological analyses and syringe tests proved the injectability of the hydrogel systems. Release studies indicated that Td and Mel were released from the fibers inside the hydrogels sequentially due to their specific locations. This release pattern from the hydrogels resulted in DPSC proliferation and odontogenic differentiation in vitro. Gene expression studies showed that the upregulation of DMP1, DSPP, and Axin-2 genes was promoted by GelMA/PecTH carrying PMMA/SF loaded with Mel (50 µg/mL) and Td (200 nM), respectively. Our results suggest that this hydrogel system holds promise for use in the regeneration of pulp tissue.

Magnetic chitosan-silk fibroin hydrogel/graphene oxide nanobiocomposite for biological and hyperthermia applications.

This work represents a biocompatible magnetic nanobiocomposite prepared by the composition of chitosan (CS) hydrogel, silk fibroin (SF), graphene oxide (GO), and Fe3O4 NPs. Terephthaloyl thiourea was applied as a cross-linking agent to cross-link the CS strings. The CS hydrogel/SF/GO/Fe3O4 nanobiocomposite with many characteristics, such as high structural uniformity, thermal stability, biocompatibility, and stability in an aqueous solution. Various characteristics of this novel magnetic nanobiocomposite were distinguished by FT-IR, EDX, FE-SEM, XRD, TGA, and VSM analysis. The FE-SEM images were taken to evaluate the size distribution of the magnetic nanoparticles (MNPs) between 39.9 and 73.3 nm as well. The performance of the prepared nanobiocomposite was assessed by the magnetic fluid hyperthermia process. Under the alternating magnetic field (AMF), the mean value of the specific absorption rate (SAR) was determined at 43.15 w/g.

Biological investigation of a novel nanocomposite based on functionalized graphene oxide nanosheets with pectin, silk fibroin and zinc chromite nanoparticles.

For biotechnology applications, a novel nanobiocomposite was synthesized based on modification of graphene oxide (GO) by extracted silk fibroin (SF), natural polymer pectin (Pec) and zinc chromite (ZnCr2O4) nanoparticles (NPs). The structure and properties of hybrid nanobiocomposite GO-Pec/SF/ZnCr2O4 such as thermal stability, less toxicity, biocompatibility, antibacterial, and biodegradable were proved by using field emission scanning electron microscope (FE-SEM), Fourier-transformed infrared (FT-IR), Energy dispersive X-ray spectroscopy (EDS), thermal gravimetric analysis (TGA), and X-Ray diffraction (XRD). According to the biological features of substances, the GO-Pec/SF/ZnCr2O4 nanobiocomposite shows perfect results in MTT (83.71 %) and Hemolysis (16.52 %) assays. accordingly, mentioned properties of this nanobiocomposite can be used as a scaffold for medical applications.

Silk protein/polyvinylpyrrolidone nanofiber membranes loaded with puerarin accelerate wound healing in mice by reducing the inflammatory response.

In modern clinical applications, wound healing remains a considerable challenge. Excessive inflammatory response is associated with delayed wound healing. In this study, we prepared composite nanofibrous membranes by mixing the Chinese herbal extract puerarin (PUE) with natural silk protein (SF) and synthetic polymer polyvinylpyrrolidone (PVP) using electrostatic spinning technique, and conducted a series of studies on the structural and biological properties of the fibrous membranes. The results showed that the loading of PUE increased the diameter, porosity and hydrophilicity of nanofibers, which were more favorable for cell adhesion and proliferation. ABTS radical scavenging assay also showed that the loading of PUE enhanced the antioxidant properties of the fibrous membranes. In addition, SF/PVP/PUE nanofibers are non-toxic and can be used as wound dressings. In vitro experiments showed that SF/PVP/PUE nanofibers could effectively alleviate lipopolysaccharide (LPS)-induced inflammation in Immortalized human keratinocytes (HaCaT) cells and down-regulate pro-inflammatory cytokine expression in cells. In vivo studies further showed that the SF/PVP/PUE nanofibers could effectively accelerate wound repair. The mechanism is that SF/PVP/PUE nanofibers can inhibit the activation and transduction of toll-like receptor 4/myeloid differentiation factor88/nuclear factor kappa B (TLR4/MyD88/NF-κB) and phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) signaling pathways, thereby reducing the inflammatory response and achieving wound healing.

Silk fibroin hydrogel containing Sesbania sesban L. extract for rheumatoid arthritis treatment.

PURPOSE: Rheumatoid arthritis, a chronic and progressive inflammation condition in the joints, has significantly reduced the patient quality of life and life expectancy. Crucially, there is no complete therapy for this disease, and the current treatments possess numerous side effects. Thus, novel therapeutic approach is necessary. To that end, this study developed novel silk fibroin in-situ hydrogel containing Sesbania sesban L. extract, a plant with high anti-inflammatory actions that are beneficial for rheumatoid arthritis treatments. METHODS: The hydrogels were manufactured using simple method of spontaneous gelation at different temperature. The gel properties of morphology, gelation time, viscosity, gel strength, stability, drug loading capacity, drug release rate, and in-vitro anti-inflammatory activity were investigated with appropriate methods. RESULTS: The optimal formulation had highly porous structure, with a gelation time of 0.5 h at room temperature and bodily temperature of 37 °C, a viscosity of 2530 ± 50 cP, a gel strength of 1880.14 ± 35.10 g, and a physical stability of >6 months. Moreover, the hydrogel contained the Sesbania sesban L. leaf extract with a total phenolic content of 92.8 ± 8.30 mg GAE/g, and sustained the release rate for >20 days, followed the Higuchi model. Regarding the in-vitro activities, all formulations were nontoxic to the RAW 264.7 cell line and demonstrated comparable anti-inflammatory activity to the free extract, in terms of the NO reduction levels. CONCLUSION: Conclusively, the systems possessed potential properties to be further investigated to become a prospective rheumatoid arthritis treatment.

Fabrication and biomedical applications of Arabinoxylan, Pectin, Chitosan, soy protein, and silk fibroin hydrogels via laccase - Ferulic acid redox chemistry.

The unique physiochemical properties and the porous network architecture of hydrogel seek the attention to be explored in broad range of fields. In the last decade, numerous studies on the development of enzymatically cross-linked hydrogels have been elucidated. Implementing enzyme based cross-linking for fabrication of biomaterials over other cross-linking methods harbor various advantages, especially hydrogels designed using laccase exhibits mild reaction environment, high cross-linking efficiency and less toxicity. To our knowledge this is the first report reviewing the formulation of laccase mediated cross-linking for hydrogel preparation. Here, laccase catalyzed synthesis of hydrogel using polysaccharide viz. arabinoxylan, sugar beet pectin, galactomannan, chitosan etc. and proteins namely soy protein, gelatin, silk fibroin were discussed on highlighting their mechanical properties and its possible field of application. We have summarized the role of phenolic acids in laccase mediated cross-linking particularly ferulic acid which is a component of lignocellulose, serving cell rigidity via cross-linkage. The review also discusses on various biomedical applications such as controlled protein release, tissue engineering, and wound healing. It is anticipated that this review will give a detailed information on different laccase mediated reaction strategies that can be applied for the synthesis of various new biomaterials with tailor made properties.

Pectin-cellulose hydrogel, silk fibroin and magnesium hydroxide nanoparticles hybrid nanocomposites for biomedical applications.

Natural polymers are at the center of materials development for biomedical and biotechnological applications based on their biocompatibility, low-toxicity and biodegradability. In this study, a novel nanobiocomposite based on cross-linked pectin-cellulose hydrogel, silk fibroin, and Mg(OH)2 nanoparticles was designed and synthesized. After extensive physical-chemical characterization, the biological response of pectin-cellulose/silk fibroin/Mg(OH)2 nanobiocomposite scaffolds was evaluated by cell viability, red blood cells hemolytic and anti-biofilm assays. After 3 days and 7 days, the cell viability of this nanobiocomposite scaffold was 65.5% and 60.5% respectively. The hemolytic effect was below 20%. Furthermore, the presence of silk fibroin and Mg(OH)2 nanoparticles allowed to enhance the anti-biofilm activity, inhibiting the P. aeruginosa biofilm formation.

Egg Case Protein 3: A Constituent of Black Widow Spider Tubuliform Silk.

Spider silk has outstanding mechanical properties, rivaling some of the best materials on the planet. Biochemical analyses of tubuliform silk have led to the identification of TuSp1, egg case protein 1, and egg case protein 2. TuSp1 belongs to the spidroin superfamily, containing a non-repetitive N- and C-terminal domain and internal block repeats. ECP1 and ECP2, which lack internal block repeats and sequence similarities to the highly conserved N- and C-terminal domains of spidroins, have cysteine-rich N-terminal domains. In this study, we performed an in-depth proteomic analysis of tubuliform glands, spinning dope, and egg sacs, which led to the identification of a novel molecular constituent of black widow tubuliform silk, referred to as egg case protein 3 or ECP3. Analysis of the translated ECP3 cDNA predicts a low molecular weight protein of 11.8 kDa. Real-time reverse transcription-quantitative PCR analysis performed with different silk-producing glands revealed ECP3 mRNA is predominantly expressed within tubuliform glands of spiders. Taken together, these findings reveal a novel protein that is secreted into black widow spider tubuliform silk.

Selenium nanoparticles coated with pH responsive silk fibroin complex for fingolimod release and enhanced targeting in thyroid cancer.

Cancer-targeted drug delivery systems based on nanoparticles (NPs) have been considered promising therapies. In this study, we developed a pH-responsive smart NPs drug delivery system using silk fibroin (SF), selenium nanoparticles (Se NPs), fingolimod (FTY720), and heptapeptide (T7). The prepared FTY720@T7-SF-Se NPs were spheres with an average diameter of 120 nm, which would contribute to the enhanced permeability and retention effects in tumour regions. The encapsulation efficiency (EE) of the FTY720@T7-SF-Se NPs was 71.95 ± 3.81%. The release of FTY720 from the nanocarriers was pH-dependent, and the release of FTY720 was accelerated in an acidic environment. Both in vitro and in vivo studies showed that FTY720@T7-SF-Se NPs had an enhanced cellular uptake selectivity and antitumor activity for thyroid cancer. The bio-distribution study in vivo further demonstrated that FTY720@T7-SF-Se NPs could effectively accumulate in the tumour region, thereby enhancing the ability to kill cancer cells in vivo. In addition, studies of histology and immunohistochemistry showed that FTY720@T7-SF-Se NPs had low toxicity to the major organs of tumour-bearing mice, indicating the prepared NPs has good biocompatibility in vivo. These results suggest that the tumour-targeted NPs delivery system (FTY720@T7-SF-Se NPs) has great potential as a new tool for thyroid cancer therapy.

Engineering luminescent pectin-based hydrogel for highly efficient multiple sensing.

Luminescent hydrogels with sensing capabilities have attracted much interest in recent years, especially those responsive to stimuli, making such materials potential for various applications. Pectin is a high-molecular-weight carbohydrate polymer that has the ability to form hydrogel upon heating or mixing with divalent cations. However, intrinsic pectin gels are weak and lack of functionalities. In this study, lanthanide ions and silk fibroin derived carbon dots were incorporated into Pectin/PVA hydrogel (PPH) to form luminescent tough hydrogels. The luminescence of the hydrogel can be tuned by adjusting the ratio of blue emission carbon dots to Eu3+ ions (red emission) and Tb3+ ions (green emission). Such incorporation of emitters only slightly changed the mechanical properties of the tough hydrogel. Notably, the luminescent Pectin/PVA hydrogel (LPPH) showed chromic response to external stimuli, like pH and metal ions. By measuring the ratio of luminescent intensity at 473 nm and 617 nm (I473/I617), the pH response can be quantified in high sensitivity. In addition, the specific detection of Cu2+ and Fe3+ ions using the fabricated hydrogel were demonstrated, the mechanism was also proposed. The different chromic responses to Fe2+ and Fe3+ endow the luminescent tough Pectin/PVA hydrogel potential for multiple sensing applications.

Chondrogenic differentiation of Wharton's Jelly mesenchymal stem cells on silk spidroin-fibroin mix scaffold supplemented with L-ascorbic acid and platelet rich plasma.

In this research, hWJ-MSCs were grown on silk scaffolds and induced towards chondrogenesis by supplementation with L-ascorbic acid (LAA) or platelet rich plasma (PRP). Silk scaffolds were fabricated with salt leaching method by mixing silk fibroin (SF) with silk spidroin (SS). The silk fibroin was obtained from Bombyx mori cocoon that had been degummed, and the silk spidroin was obtained from wild-type spider Argiope appensa. The effect of scaffold composition and inducer on cell proliferation was observed through MTT assay. The most optimal treatment then continued to be used to induce hWJ-MSC towards chondrogenic differentiation for 7 and 21 days. Scaffolds characterization showed that the scaffolds produced had 3D structure with interconnected pores, and all were biocompatible with hWJ-MSCs. Scaffold with the addition of 10% SS + 90% SF showed higher compressive strength and better pore interconnectivity in comparison to 100% silk fibroin scaffold. After 48 h, cells seeded on scaffold with spidroin and fibroin mix had flattened morphology in comparison to silk fibroin scaffold which appeared to be more rounded on the scaffold surface. Scaffold with 10% (w/w) of silk spidroin (SS) + 90% (w/w) of silk fibroin (SF) was the most optimal composition for cell proliferation. Immunocytochemistry of integrin ß1 and RGD sequence, showed that scaffold with SS 10% provide better cell attachment with the presence of RGD sequence from the spidroin silk which could explain the higher cell proliferation than SF100% scaffold. Based on Alcian Blue staining and Collagen Type II immunocytochemistry (ICC), cells grown on 10% SS + 90% SF scaffold with 10% PRP supplementation were the most optimal to support chondrogenesis of hWJ-MSCs. These results showed that the addition of spidroin silk from A. appensa. had impact on scaffold compressive strength and chondrogenic differentiation of hWJ-MSC and had the potential for further development of bio-based material scaffold in cartilage tissue engineering.

Oral Bioavailability Evaluation of Celastrol-Encapsulated Silk Fibroin Nanoparticles Using an Optimized LC-MS/MS Method.

Celastrol (CL), a compound isolated from Tripterygium wilfordii, possesses various bioactivities such as antitumor, anti-inflammatory and anti-obesity effects. In previous studies, we developed CL-encapsulated silk fibroin nanoparticles (CL-SFNP) with satisfactory formulation properties and in vitro cancer cytotoxicity effect. For further in vivo oral bioavailability evaluation, in this study, a simple and reliable LC-MS/MS method was optimized and validated to determine CL concentration in rat plasma. The separation of CL was performed on a C18 column (150 by 2 mm, 5 µm) following sample preparation using liquid-liquid extraction with the optimized extraction solvent of tert-butyl methylether. The assay exhibited a good linearity in the concentration range of 0.5-500 ng/mL with the lower limit of quantification (LLOQ) of 0.5 ng/mL. The method was validated to meet the requirements for bioassay with accuracy of 91.1-110.0%, precision (RSD%) less than 9.1%, extraction recovery of 63.5-74.7% and matrix effect of 87.3-101.2%. The developed method was successfully applied to the oral bioavailability evaluation of CL-SFNP. The pharmacokinetic results indicated the AUC0-∞ values of CL were both significantly (p < 0.05) higher than those for pure CL after intravenous (IV) or oral (PO) administration of equivalent CL in rats. The oral absolute bioavailability (F, %) of CL significantly (p < 0.05) increased from 3.14% for pure CL to 7.56% for CL-SFNP after dosage normalization. This study provides valuable information for future CL product development.