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The purpose of this work was to investigate the feasibility of a novel ophthalmic formulation of amphotericin B-encapsulated silk fibroin nanoparticles incorporated in situ hydrogel (AmB-FNPs ISG) for fungal keratitis (FK) treatment. AmB-FNPs ISG composites were successfully developed and have shown optimized physicochemical properties for ocular drug delivery. Antifungal effects against Candida albicans and in vitro ocular irritation using corneal epithelial cells were performed to evaluate the efficacy and safety of the composite formulations. The combined system of AmB-FNPs-ISG exhibited effective antifungal activity and showed significantly less toxicity to HCE cells than commercial AmB. In vitro and ex vivo mucoadhesive tests demonstrated that the combination of silk fibroin nanoparticles with in situ hydrogels could enhance the adhesion ability of the particles on the ocular surface for more than 6 h, which would increase the ocular retention time of AmB and reduce the frequency of administration during the treatment. In addition, AmB-FNP-PEG ISG showed good physical and chemical stability under storage condition for 90 days. These findings indicate that AmB-FNP-PEG ISG has a great potential and be used in mucoadhesive AmB eye drops for FK treatment. [ABSTRACT FROM AUTHOR]

Abstract Background Reduced supplies of oxygen and nutrients caused by vascular injury lead to difficult-to-heal pressure ulcers (PU) in clinical practice. Rapid vascular repair in the skin wound is the key to the resolution of this challenge, but clinical measures are still limited. We described the beneficial effects of extracellular vesicle-derived silk fibroin nanoparticles (NPs) loaded with milk fat globule EGF factor 8 (MFGE8) on accelerating skin blood vessel and PU healing by targeting CD13 in the vascular endothelial cells (VECs). Methods CD13, the specific targeting protein of NGR, and MFGE8, an inhibitor of ferroptosis, were detected in VECs and PU tissues. Then, NPs were synthesized via silk fibroin, and MFGE8-coated NPs (NPs@MFGE8) were assembled via loading purified protein MFGE8 produced by Chinese hamster ovary cells. Lentivirus was used to over-express MFGE8 in VECs and obtained MFGE8-engineered extracellular vesicles (EVs-MFGE8) secreted by these VECs. The inhibitory effect of EVs-MFGE8 or NPs@MFGE8 on ferroptosis was detected in vitro. The NGR peptide cross-linked with NPs@MFGE8 was assembled into NGR-NPs@MFGE8. Collagen and silk fibroin were used to synthesize the silk fibroin/collagen hydrogel. After being loaded with NGR-NPs@MFGE8, silk fibroin/collagen hydrogel sustained-release carrier was synthesized to investigate the repair effect on PU in vivo. Results MFGE8 was decreased, and CD13 was increased in PU tissues. Similar to the effect of EVs-MFGE8 on inhibiting ferroptosis, NPs@MFGE8 could inhibit the mitochondrial autophagy-induced ferroptosis of VECs. Compared with the hydrogels loaded with NPs or NPs@MFGE8, the hydrogels loaded with NGR-NPs@MFGE8 consistently released NGR-NPs@MFGE8 targeting CD13 in VECs, thereby inhibiting mitochondrial autophagy and ferroptosis caused by hypoxia and accelerating wound healing effectively in rats. Conclusions The silk fibroin/collagen hydrogel sustained-release carrier loaded with NGR-NPs@MFGE8 was of great significance to use as a wound dressing to inhibit the ferroptosis of VECs by targeting CD13 in PU tissues, preventing PU formation and promoting wound healing. Graphical Abstract

Background: Reduced supplies of oxygen and nutrients caused by vascular injury lead to difficult-to-heal pressure ulcers (PU) in clinical practice. Rapid vascular repair in the skin wound is the key to the resolution of this challenge, but clinical measures are still limited. We described the beneficial effects of extracellular vesicle-derived silk fibroin nanoparticles (NPs) loaded with milk fat globule EGF factor 8 (MFGE8) on accelerating skin blood vessel and PU healing by targeting CD13 in the vascular endothelial cells (VECs). Methods: CD13, the specific targeting protein of NGR, and MFGE8, an inhibitor of ferroptosis, were detected in VECs and PU tissues. Then, NPs were synthesized via silk fibroin, and MFGE8-coated NPs (NPs@MFGE8) were assembled via loading purified protein MFGE8 produced by Chinese hamster ovary cells. Lentivirus was used to over-express MFGE8 in VECs and obtained MFGE8-engineered extracellular vesicles (EVs-MFGE8) secreted by these VECs. The inhibitory effect of EVs-MFGE8 or NPs@MFGE8 on ferroptosis was detected in vitro. The NGR peptide cross-linked with NPs@MFGE8 was assembled into NGR-NPs@MFGE8. Collagen and silk fibroin were used to synthesize the silk fibroin/collagen hydrogel. After being loaded with NGR-NPs@MFGE8, silk fibroin/collagen hydrogel sustained-release carrier was synthesized to investigate the repair effect on PU in vivo. Results: MFGE8 was decreased, and CD13 was increased in PU tissues. Similar to the effect of EVs-MFGE8 on inhibiting ferroptosis, NPs@MFGE8 could inhibit the mitochondrial autophagy-induced ferroptosis of VECs. Compared with the hydrogels loaded with NPs or NPs@MFGE8, the hydrogels loaded with NGR-NPs@MFGE8 consistently released NGR-NPs@MFGE8 targeting CD13 in VECs, thereby inhibiting mitochondrial autophagy and ferroptosis caused by hypoxia and accelerating wound healing effectively in rats. Conclusions: The silk fibroin/collagen hydrogel sustained-release carrier loaded with NGR-NPs@MFGE8 was of great significance to use as a wound dressing to inhibit the ferroptosis of VECs by targeting CD13 in PU tissues, preventing PU formation and promoting wound healing. [ABSTRACT FROM AUTHOR]

Simple Summary: For many types of cancer, chemotherapy is still widely used, but it can cause serious side effects because it kills all rapidly growing cells, not just the cancerous ones. In this paper, a nanoparticle-based drug delivery system has been designed to selectively deliver a highly cytotoxic drug to tumor cells. The nanoparticles, obtained from a protein called silk fibroin, have been loaded with a naphthalene diimide derivative, namely NDI-1, which kills cells by affecting their DNA. The surface of the silk fibroin nanoparticles has been decorated with cyclopentapeptides incorporating the Arg-Gly-Asp sequence (cRGDs) to specifically target the tumor cells, thus reducing the harm to healthy cells and minimizing side effects. Developing drug delivery systems to target cytotoxic drugs directly into tumor cells is still a compelling need with regard to reducing side effects and improving the efficacy of cancer chemotherapy. In this work, silk fibroin nanoparticles (SFNs) have been designed to load a previously described cytotoxic compound (NDI-1) that disrupts the cell cycle by specifically interacting with non-canonical secondary structures of DNA. SFNs were then functionalized on their surface with cyclic pentapeptides incorporating the Arg-Gly-Asp sequence (cRGDs) to provide active targeting toward glioma cell lines that abundantly express ανβ3 and ανβ5 integrin receptors. Cytotoxicity and selective targeting were assessed by in vitro tests on human glioma cell lines U373 (highly-expressing integrin subunits) and D384 cell lines (low-expressing integrin subunits in comparison to U373). SFNs were of nanometric size (d50 less than 100 nm), round shaped with a smooth surface, and with a negative surface charge; overall, these characteristics made them very likely to be taken up by cells. The active NDI-1 was loaded into SFNs with high encapsulation efficiency and was not released before the internalization and degradation by cells. Functionalization with cRGDs provided selectivity in cell uptake and thus cytotoxicity, with a significantly higher cytotoxic effect of NDI-1 delivered by cRGD-SFNs on U373 cells than on D384 cells. This manuscript provides an in vitro proof-of-concept of cRGD-silk fibroin nanoparticles' active site-specific targeting of tumors, paving the way for further in vivo efficacy tests. [ABSTRACT FROM AUTHOR]

Silk fibroin nanoparticles (SFNs) have been widely investigated for drug delivery, but their clinical application still faces technical (large-scale and GMP-compliant manufacturing), economic (cost-effectiveness in comparison to other polymer-based nanoparticles), and biological (biodistribution assessments) challenges. To address biodistribution challenge, we provide a straightforward desolvation method (in acetone) to produce homogeneous SFNs incorporating increasing amounts of Fe 2 O 3 (SFNs-Fe), detectable by Magnetic Resonance Imaging (MRI), and loaded with curcumin as a model lipophilic drug. SFNs-Fe were characterized by a homogeneous distribution of the combined materials and showed an actual Fe 2 O 3 loading close to the theoretical one. The amount of Fe 2 O 3 incorporated affected the physical-chemical properties of SFNs-Fe, such as polymer matrix compactness, mean diameter and drug release mechanism. All formulations were cytocompatible; curcumin encapsulation mitigated its cytotoxicity, and iron oxide incorporation did not impact cell metabolic activity but affected cellular uptake in vitro. SFNs-Fe proved optimal for biodistribution studies, as MRI showed significant nanoparticle retention at the administration site, supporting their potential for locoregional cancer therapy. Finally, technical and economic challenges in SFN production were overcome using a GMP-compliant microfluidic scalable technology, which optimized preparation to produce smaller particle sizes compared to manual methods and reduced acetone usage, thus offering environmental and economic benefits. Moreover, enabling large-scale production of GMP-like SFNs, this represents a considerable step forward for their application in the clinic. • SFNs face challenges in large-scale manufacturing, cost-effectiveness and biodistribution assessments. • SFNs incorporating increasing amounts of Fe 2 O 3 , detectable by MRI, and loaded with curcumin as a model drug were prepared. • The amount of Fe 2 O 3 incorporated affected polymer matrix compactness, mean diameter and curcumin release through diffusion. • MRI showed SFNs-Fe retention at the administration site, supporting their potential for locoregional cancer therapy. • GMP-compliant microfluidic optimized preparation even reducing acetone usage, offering environmental and economic benefits. [ABSTRACT FROM AUTHOR]

Instability of silk fibroin nanoparticles (SFNPs) in physiologic condition hinders its application as drug delivery vehicle. Herein, indocyanine green (ICG) loaded silk fibroin nanoparticles (ICG-SFNPs) was firstly prepared and then crosslinked by proanthocyanidins to obtain the stable ICG-CSFNPs for killing the residual tumour niche under near infra-red irradiation (NIR) after surgery. The particle size and zeta potentials of ICG-CSFNPs was 120.1 nm and -40.4 mV, respectively. Moreover, ICG-CSFNPs exhibited good stability of particle size in the physiological medium. Meanwhile, the stable photothermal properties of ICG-CSFNPs were not compromised even after several cycles of NIR. Few of the ICG-CSFNPs were phagocytized by RAW264.7 macrophage in vitro, while they were easily internalized by C6 glioma cells, resulting in their significant toxicity on tumour cells after NIR. The pharmacokinetic study showed that ICG-CSFNPs had a longer blood circulation time than ICG-SFNPs, making them more distribution in glioma after intravenous administration in vivo. Meanwhile, the pharmacological study showed the more effective inhibition of tumour growth was exhibited by ICG-CSFNPs in C6 glioma-bearing mice after NIR. Overall, the cross-linked nanoparticles of silk fibroin may be a promising vehicle of ICG for photothermal therapy of glioma after surgical resection.

Silk fibroin nanoparticles possess the hydrophobic nature which assists them to become a good substrate for reticulo-endothelial system (RES) and macrophageal uptake. Surface coating of these nanoparticles with hydrophilic stabilizers, like Tween-80 make them long circulating and facilitate their uptake by low density lipoprotein (LDL) receptors to cross blood brain barrier (BBB). Surface modified silk fibroin nanoparticles bearing anti-cancer agent doxorubicin (DOX) were fabricated by desolvation method and coated with Tween-80 as surface modifier. The prepared nanoparticles were characterized for various physicochemical parameters, like particle size, surface charge, surface morphology by scanning electron microscope (SEM) and transmission electron microscopy (TEM), and in vitro drug release along with in vitro cell cytotoxicity, flow cytometry and cellular uptake studies by flourocytometry on glioblastoma cell lines. Entrapment efficiency for the silk fibroin nanoparticles were found to be >85% for coated and uncoated nanoparticles. Nanoparticles with average diameter less than 150 nm having negative charge were found to show no toxicity of its own. The pro-inflammatory response of nanoparticles was observed by determining the cytokines level, such as TNF-α and IL-1β. Sustained drug release pattern from the nanoparticles with better cytotoxicty as compared to free drug was observed, signifying their potential ability to work as a drug delivery system. • Silk fibroin was extracted from silk worm cocoons. • Silk fibroin is used as a biological macromolecule for drug delivery. • Silk fibroin nanoparticles provide better cytotocixity effects. • Coating of nanoparticles with Tween-80 improves the nanoparticles cytotocixity effects. [ABSTRACT FROM AUTHOR]

Recently, the use of natural materials has grown in the plastics industry. In this study, novel bio-nanocomposites were developed from poly(lactic acid)/poly(ε-caprolactone) (PLA/PCL) blends and silk fibroin nanoparticles (SFNP). SFNP were successfully synthesized from silk fibroin (SF) and analyzed by Fourier-transform infrared spectroscopy (FT-IR), X-ray diffraction, thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). The obtained results revealed that the well-ordered structures in SF were changed to the amorphous structures in SFNP. Then (PLA/PCL) blends containing 10–30% of PCL were prepared and characterized. According to the obtained results, the PLA/PCL (70/30) blend was selected as the optimized sample for further studies. The scanning electron microscopy results illustrated that the addition of 1% of SFNP into this blend improved the compatibility between PLA and PCL and reduced the PCL droplet sizes from 1.170 ± 92 to 794 ± 46 nm. The results from TGA analysis indicated that the presence of SFNP enhanced the thermal stability of materials at high temperatures. The crystallization kinetics results revealed that while SFNP promoted the crystallization of neat PLA, the crystallization rate of PLA/PCL blend was decreased upon the incorporation of nanofiller. Furthermore, the PLA/PCL/SFNP exhibited higher microhardness and barrier properties than the neat blend. The results suggest that the developed bio-nanocomposites are promising materials for demanding applications such as food packaging. [ABSTRACT FROM AUTHOR]

Instability of silk fibroin nanoparticles (SFNPs) in physiologic condition hinders its application as drug delivery vehicle. Herein, indocyanine green (ICG) loaded silk fibroin nanoparticles (ICG-SFNPs) was firstly prepared and then crosslinked by proanthocyanidins to obtain the stable ICG-CSFNPs for killing the residual tumour niche under near infra-red irradiation (NIR) after surgery. The particle size and zeta potentials of ICG-CSFNPs was 120.1 nm and -40.4 mV, respectively. Moreover, ICG-CSFNPs exhibited good stability of particle size in the physiological medium. Meanwhile, the stable photothermal properties of ICG-CSFNPs were not compromised even after several cycles of NIR. Few of the ICG-CSFNPs were phagocytized by RAW264.7 macrophage in vitro, while they were easily internalized by C6 glioma cells, resulting in their significant toxicity on tumour cells after NIR. The pharmacokinetic study showed that ICG-CSFNPs had a longer blood circulation time than ICG-SFNPs, making them more distribution in glioma after intravenous administration in vivo. Meanwhile, the pharmacological study showed the more effective inhibition of tumour growth was exhibited by ICG-CSFNPs in C6 glioma-bearing mice after NIR. Overall, the cross-linked nanoparticles of silk fibroin may be a promising vehicle of ICG for photothermal therapy of glioma after surgical resection. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate intravitreal injection of silk fibroin nanoparticles (SFNs) encapsulating bio-macromolecules, achieving enhanced drug bioavailability, and extended retention in retina. SFNs were prepared with regenerated silk fibroin using desolvation method with fluorescein isothiocyanate labeled bovine serum albumin (FITC-BSA) as bio-macromolecular model drug encapsulated. In vitro physicochemical properties and in vitro drug release of FITC-BSA loaded SFNs (FITC-BSA-SFNs) were evaluated. Cytotoxicity, cellular uptake, and retention of FITC-BSA-SFNs were determined in human retinal pigment epithelial cell line (ARPE-19). In addition, in vivo distribution and safety of intravitreally administered FITC-BSA-SFNs were investigated in New Zealand white rabbits. The particle size of FITC-BSA-SFNs was 179.1 ± 3.7 nm with polydispersity index of 0.102 ± 0.033 and the zeta potential was greater than −25 mV. FITC-BSA-SFNs exhibited excellent biocompatibility with no cytotoxicity observed within 24 and 48 h in AREP-19 cells. Compared to FITC-BSA solution, FITC-BSA-SFNs showed enhanced cellular uptake and prolonged retention. Furthermore, FITC-BSA-SFNs achieved accumulated distribution and extended retention in retina in vivo following intravitreal injection compared to a single administration of free drug solution. Therefore, this bio-macromolecule delivery platform based on SFNs could have great potential in the treatment of posterior segment disorders. [ABSTRACT FROM AUTHOR]

Hui Li,1,* Jian Tian,1,2,* Anqing Wu,2 Jiamin Wang,1 Cuicui Ge,2 Ziling Sun1–3 1School of Biological and Basic Medical Science, 2School of Radiological & Interdisciplinary Sciences, Soochow University, 3Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, Suzhou, Jiangsu, People’s Republic of China *These authors contributed equally to this work Abstract: Self-assembled nanoparticles of the natural polymer, silk fibroin (SF), are a very promising candidate in drug delivery due to their biocompatible and biodegradable properties. In this study, SF nanoparticles loaded with 5-fluorouracil (5-FU) and curcumin with size 217±0.4 nm and with a loading efficacy of 45% and 15% for 5-FU and curcumin, respectively, were prepared. The in vitro release effect of 5-FU and curcumin from nanoparticles was evaluated as ~100% and ~5%, respectively. It has been revealed that the application of such a nanodrug can increase the level of reactive oxygen species, which in turn induces apoptosis of cancer cells in vitro. Animal studies have shown that tumors could be noticeably reduced after being injected with the drug-entrapped nanoparticles. More apoptotic cells were found after 7 days of treatment with SF nanoparticles by a hematoxylin–eosin staining assay. These results demonstrate the future potential of nanoparticle-loaded binary drugs in the treatment of breast cancer. Keywords: silk fibroin nanoparticles, 5-flurouracil, curcumin, reactive oxygen species

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.

Objective: the objective of the present work is to study the effectiveness of treatment with silk fibroin nanoparticles loaded with resveratrol in experimental periodontitis in a diabetic rat model. Introduction: Periodontitis is an inflammatory pathology highly related to other diseases, such as type II diabetes. Both diseases have a specific inflammatory condition, with Interleukin (IL)-6, IL-1β and Transforming Grow Factor (TGF)-1β being the most relevant proinflammatory factors. Silk fibroin (SF) nanoparticles loaded with resveratrol (Res-SFN) are a new alternative as a treatment. Methods: 40 diabetic Sprague Dawley male rats were used and periodontitis was induced by ligation. The animals were divided into 5 treatment groups, and 1 mL of treatment was administered once a day for 4 weeks. The groups were: I: Carboxymethyl cellulose (CMC) 0.8%, II: CMC 0.8% + SF 1%, III: CMC 0.8% + RES-SFN 3 mg/mL, IV: CMC 0.8% + SF 1% + RES-SFN 3 mg/mL, V: Water. A peripheral blood sample was taken every week to quantify the inflammatory profile by ELISA (IL-6, IL-1β and TGF-1β). After 4 weeks the sacrifice was carried out and biopsies of the gum were taken. Results: Treatment with SF and RES-SFN reduced the amount of chemical inflammation mediators (with the exception of IL-1β in comparisons I-IV and II-IV (p > 0.05)), as well as the anatomopathological variables linked to it, in a significant way (p < 0.05). Conclusion: treatment with RES-SFN has reduced local inflammation in this experimental periodontitis model.

Abstract: Silk fibroin nanoparticles (SFNPs) as a natural polymer have been utilized in biomedical applications such as suture, tissue engineering‐based scaffolds, and drug delivery carriers. Since there is little data regarding the toxicity effects on different cells and tissues, we aimed to determine the toxicity mechanisms of SFNPs on human lymphocytes and monocytes based on reliable methods. Our results showed that SFNPs (0.5, 1, and 2 mg/mL) induced oxidative stress via increasing reactive oxygen species production, mitochondrial membrane potential (∆Ψ) collapse, which was correlated to cytochrome c release and Adenosine diphosphate (ADP)/Adenosine tri phosphate (ATP) ratio increase as well as lysosomal as another toxicity mechanism, which led to cytosolic release of lysosomal digestive proteases, phosphor lipases, and apoptosis signaling. Taken together, these data suggested that SFNPs toxicity was associated with mutual mitochondrial/lysosomal cross‐talk and oxidative stress on human lymphocytes and monocytes with activated apoptosis signaling. [ABSTRACT FROM AUTHOR]

Celastrol (CL), a bioactive compound isolated from Tripterygium wilfordii, has demonstrated bioactivities against a variety of diseases including cancer and obesity. However, its poor water solubility and rapid in vivo clearance limit its clinical applications. To overcome these limitations, nanotechnology has been employed to improve its pharmacokinetic properties. Nanoparticles made of biological materials offer minimal adverse effects while maintaining the efficacy of encapsulated therapeutics. Silk fibroin (SF) solution was prepared successfully by extraction from the cocoons of silkworms, and a final concentration of 2 mg/mL SF solution was used for the preparation of CL-loaded SF nanoparticles (CL-SFNP) by the desolvation method. A stirring speed of 750 rpm and storage time of 20 h at −20 °C resulted in optimized product yield. A high-performance liquid chromatography (HPLC) method was developed and validated for the analysis of CL in rat plasma in terms of selectivity, linearity, intra-/inter-day precision and accuracy, and recovery. No interference was observed in rat plasma. Linearity in the concentration range of 0.05−5 µg/mL was observed with R2 of 0.999. Precision and accuracy values were below the limit of acceptance criteria, i.e., 15% for quality control (QC) samples and 20% for lower limit of quantification (LLOQ) samples. Rats were given intravenous (IV) administration of 1 mg/kg of pure CL in PEG 300 solution or CL-SFNP. The pharmacokinetic profile was improved with CL-SFNP compared to pure CL. Pure CL resulted in a maximum concentration (Cmax) value of 0.17 µg mL−1 at 5 min following administration, whereas that for CL-SFNP was 0.87 µg mL−1 and the extrapolated initial concentrations (C0) were 0.25 and 1.09 µg mL−1, respectively, for pure CL and CL-SFNP. A 2.4-fold increase in total area under the curve (AUC0-inf) (µg h mL−1) was observed with CL-SFNP when compared with pure CL. CL-SFNP demonstrated longer mean residence time (MRT; 0.67 h) than pure CL (0.26 h). In conclusion, the preparation of CL-SFNP was optimized and the formulation demonstrated improved pharmacokinetic properties compared to CL in solution following IV administration.

Due to its unique properties, bacterial cellulose (BC) has attracted a great deal of interest as an implant material for tissue regeneration. However, one major problem of BC is inadequate vascularization which leads to cell apoptosis due to insufficient nutrients and oxygen supply. Herein, porous BC/gelatin (BC/Gel) scaffolds loaded with vascular endothelial growth factor (VEGF) with silk fibroin nanoparticles (VEGF-NPs) were prepared. An in vitro study indicated that VEGF was sustainably released from the BC/Gel/VEGF-NPs scaffold over 28 days. Cell viability, morphology and proliferation were evaluated using Live/Dead viability/cytotoxicity assay, field emission scanning electron microscopy and CCK-8 assay by seeding the scaffolds with pig iliac endothelium cells. The presence of VEGF-NPs in the scaffold significantly improved cell proliferation and viability in vitro. Evaluation of in vivo biocompatibility and angiogenesis of the BC/Gel/VEGF-NPs scaffold was conducted using a dog skin defect model. Results indicated that the BC/Gel/VEGF-NPs scaffold significantly promoted vessel blood formation after implantation compared to the BC/Gel and BC/Gel/NPs scaffolds. It is concluded that angiogenesis could be improved through the incorporation of VEGF-NPs into the BC/Gel scaffold, which may enhance clinically desirable functions of BC-based scaffolds in terms of enhanced angiogenesis. [ABSTRACT FROM AUTHOR]

Chitosan is a biocompatible polysaccharide with positive Z potential which can stabilize negative charged nanoparticles. Silk fibroin nanoparticles and citrate gold nanoparticles, both with negative Z potential, but they form aggregates at physiological ionic strength. In this work, we study the behavior of chitosan in solution when the ionic strength of the medium is increased and how the concentration of chitosan and the proportion of the two components (chitosan and AuNP or SFN) significantly affect the stability and size of the nanocomposites formed. In addition to experimental measurements, molecular modeling were used to gain insight into how chitosan interacts with silk fibroin monomers, and to identify the main energetic interactions involved in the process. The optimum values for obtaining the smallest and most homogeneous stable nanocomposites were obtained and two different ways of organization through which chitosan may exert its stabilizing effect were suggested. [ABSTRACT FROM AUTHOR]

Silk fibroin nanoparticles (SFN) are interesting drug delivery systems due to their high load capacity and low toxicity. The aim of this study was to reveal how SFN load red wine polyphenols (RWP), and to assess their protective effect during the early stages of digestion by using the dynamic gastrointestinal simulator (simgi®) under physiological conditions. RWP-SFN were prepared by incubation of SFN in red wine and recovered by centrifugation. The obtained nanoparticles (particle size of 194.4 ± 2.1 nm and a Zeta potential of −22.7 ± 0.1 mV) showed a significant polyphenol loading content (7386.59 ± 1018.28 μg/g) with a high encapsulation efficiency (66.66 ± 2.58%). They were highly stable during their passage through the digestive tract, being the activity of RWP highly preserved. This data paves the way for future developments of SFN as stabilizers with a wide range of applications in the field of food additives. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa (P. aeruginosa) causing urinary tract infections (UTIs) are a major concern among hospital-acquired infections. The need for an effective vaccine that reduces the infections is imperative. This study aims to evaluate the efficacy of a multi-epitope vaccine encapsulated in silk fibroin nanoparticles (SFNPs) against P. aeruginosa-mediated UTIs. A multi-epitope is constructed from nine proteins of P. aeruginosa using immunoinformatic analysis, expressed, and purified in BL21 (DE3) cells. The encapsulation efficiency of the multi-epitope in SFNPs is 85% with a mean particle size of 130 nm and 24% of the encapsulated antigen is released after 35 days. The vaccine formulations adjuvanted with SFNPs or alum significantly improve systemic and mucosal humoral responses and the cytokine profile (IFN-γ, IL-4, and IL-17) in mice. Additionally, the longevity of the IgG response is maintained for at least 110 days in a steady state. In a bladder challenge, mice treated with the multi-epitope admixed with alum or encapsulated in SFNPs demonstrate significant protection of the bladder and kidneys against P. aeruginosa. This study highlights the promising therapeutic potential of a multi-epitope vaccine encapsulated in SFNPs or adjuvanted with alum against P. aeruginosa infections., (© 2023 Wiley-VCH GmbH.)

Cervical diseases such as lymph node disease and tubal obstruction have threatened women's health. However, the traditional diagnostic methods still have shortcomings. NIR-II fluorescence imaging with advantages of low scattering, negligible autofluorescence, and high spatial resolution could be an ideal option. To obtain high quality NIR-II fluorescence imaging, selecting appropriate nanoprobes becomes the important issue. As a small molecular photothermal agent, extensive applications of ICG are rather limited because of its drawbacks. Herein, natural silk fibroin (SF) was synthesized and encapsulated ICG molecules to form SF@ICG nanoparticles (NPs). After detailed analysis, SF@ICG NPs showed excellent stability and long circulation time, as well as strong NIR-II fluorescence emission, well photo-stability, biocompatibility and well photothermal property under 808 nm laser irradiation. Furthermore, SF@ICG NPs were utilized for NIR-II fluorescence imaging of lymph node/lymphangiography and angiography of fallopian tubes. The process of fallopian tubes could be detected with high resolution and high sensitivity. A novel type of SF@ICG NPs were designed and synthesized with high efficiency. SF@ICG NPs obtained well photothermal property, biocompatibility, low cytotoxicity, strong NIR-II fluorescence intensity. Thus, SF@ICG NPs were further utilized forNIR-II fluorescence imaging of cervical disease. High quality in vivo NIR-II fluorescence imaging of sentinel lymph node (SLN) and anatomy structure of lymph-vessel were achieved with assistance of NIR-II fluorescence imaging and SF@ICG NPs. Furthermore, NIR-II fluorescence hysterography of SF@ICG NPs showed preferable performance in outlining the uteruses and natural peristalsis. [Display omitted] [ABSTRACT FROM AUTHOR]

This work aimed to establish a simple and feasible method to obtain silk fibroin nanoparticles (SFNPs) with uniform particles size, and then modify the SFNPs with nanobody (Nb) 11C12 targeting the proximal membrane end of carcinoembryonic antigen on the surface of colorectal cancer (CRC) cells. The regenerated silk fibroin (SF) was isolated using ultrafiltration tubes with a 50 kDa molecular weight cut-off, and the retention fraction (named as SF > 50 kDa) was further self-assembled into SFNPs by ethanol induction. Scanning electron microscope (SEM) and high-resolution transmission electron microscop showed that the SFNPs with uniform particles size were formed. Due to electrostatic adsorption and pH responsiveness, SFNPs have been proved to effectively load and release the anticancer drug doxorubicin hydrochloride (DOX) (DOX@SFNPs). Further, targeting molecule Nb 11C12 was used to modify these nanoparticles, constituting the targeted outer layer of the drug delivery system (DOX@SFNPs-11C12), achieving precise localization to cancer cells. The release amount of DOX observed from in vitro drug release profiles increased as follows: pH 7.4 < pH 6.8 < pH 5.4, demonstrating that the DOX release could be accelerated in a weakly acidic environment. In vitro cytotoxicity experiments displayed that SFNPs-11C12 nanoparticles exhibited good safety and biocompatibility. Drug-loaded nanoparticles, DOX@SFNPs-11C12, led to higher LoVo cells apoptosis compared to DOX@SFNPs. Fluorescence spectrophotometer characterization and confocal laser scanning microscopy further showed that the internalization of DOX was highest in the DOX@SFNPs-11C12, certifying that the introduced targeting molecule enhanced the uptake of drug delivery system by LoVo cells. This study provides a simple and operational approach to developing an optimized SFNPs drug delivery system modified by targeting Nb, which can be a good candidate for CRC therapy., (© 2023 IOP Publishing Ltd.)

Cervical diseases such as lymph node disease and tubal obstruction have threatened women's health. However, the traditional diagnostic methods still have shortcomings. NIR-II fluorescence imaging with advantages of low scattering, negligible autofluorescence, and high spatial resolution could be an ideal option. To obtain high quality NIR-II fluorescence imaging, selecting appropriate nanoprobes becomes the important issue. As a small molecular photothermal agent, extensive applications of ICG are rather limited because of its drawbacks. Herein, natural silk fibroin (SF) was synthesized and encapsulated ICG molecules to form SF@ICG nanoparticles (NPs). After detailed analysis, SF@ICG NPs showed excellent stability and long circulation time, as well as strong NIR-II fluorescence emission, well photo-stability, biocompatibility and well photothermal property under 808 nm laser irradiation. Furthermore, SF@ICG NPs were utilized for NIR-II fluorescence imaging of lymph node/lymphangiography and angiography of fallopian tubes. The process of fallopian tubes could be detected with high resolution and high sensitivity., Competing Interests: Declaration of competing interest The authors declare that no competing interests exist., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

[Display omitted] Inflammatory bowel disease (IBD) is a chronic and idiopathic inflammatory disorder affecting the gastrointestinal tract. The pharmacological treatments used currently for its treatment lack efficacy, so new therapeutic strategies should be developed. In this context, flavonoids loaded in biopolymeric nanoparticles can be considered as novel promising candidates. The aim of the present study was to evaluate the intestinal anti-inflammatory effects of quercetin when is administered loaded in silk fibroin nanoparticles (QSFN) in the dextran sulphate sodium experimental model of mouse colitis, which displays some similarities to human IBD. Previously characterized quercetin-loaded silk fibroin nanoparticles (QSFN). QSFN showed a reversible aggregation profile induced by the acidification of the solution but did not affect the loaded quercetin. Daily administration of QSFN significantly reduced disease activity index values compared to the control colitic group. This beneficial effect was not only corroborated by the histological examination of the colonic specimens but also the improvement of the colonic expression of the different proinflammatory cytokines (Tnf-α , Il-1β , Il-6, Mcp-1 , Icam-1 , Nlrp3 and iNOS). Therefore, these data suggest that QSFN could be a promising alternative to current treatments as a drug delivery system for IBD treatment. [ABSTRACT FROM AUTHOR]

Nanotechnology-based approaches have enabled overcoming the challenging issues such as the rapid clearance, poor solubility, and non-specific action or cellular uptake of drugs. In this study, we have evaluated the therapeutic effects of the phenolic compound, ferulic acid (FA), in the acute pancreatitis (AP) as this phenolic compound has demonstrated promising effects against the oxidative stress and inflammatory reactions. In order to overcome the poor solubility and bioavailability of FA, it was entrapped into the nanoparticles (NPs) based on the silk fibroin (SF) as a biomimetic substance. Neutrophil membrane-coated SF-NPs with appropriate capacity of FA loading and physicochemical characteristics, released FA in a controlled fashion, selectively delivered FA into the inflammatory pancreas lesion, and demonstrated protective effects against the detrimental aspects of the disease. The prepared nanoformulation by improving the pharmacological profile of FA and targeted delivery could be of therapeutic importance against the AP via suppressing the inflammation and oxidative stress. [ABSTRACT FROM AUTHOR]

Simple Summary: Many tumor cell types overexpress integrins, a glycoprotein, on their cell membranes. The tripeptide motif Arg-Gly-Asp (RGD) is well-known for being recognized by the integrin superfamily members and can thus be used to actively target nanoparticles containing cytotoxic drugs directly to the tumor cells. According to this strategy, the antitumor activity is boosted, and healthy organs are spared. In this paper, silk fibroin, a naturally derived protein, has been used to prepare nanoparticles (SFNs) functionalized on their surface with RGD. In vitro experiments revealed that functionalization of SFNs with RGD provided active internalization by tumor cells overexpressing integrin receptors. Therefore, RGD-SFNs may be used for tumor-specific delivery of anticancer drugs. Arg-Gly-Asp (RGD)-based cyclopentapeptides (cRGDs) have a high affinity towards integrin αvβ3 and αvβ5, which are overexpressed by many tumor cells. Here, curcumin-loaded silk fibroin nanoparticles (SFNs) have been functionalized on the surface with cRGD to provide active targeting towards tumor cells; a "click reaction" between the RGD-based cyclopentapeptide carrying an azide group and triple-bond-functionalized nanoparticles has been exploited. Both naked and functionalized SFNs were less than 200 nm in diameter and showed a round-shaped morphology but, after functionalization, SFNs increased in size and protein molecular weight. The functionalization of SFNs' surfaces with cRGD provided active internalization by cells overexpressing integrin receptors. At the lowest concentration tested (0.01 mg/mL), functionalized SFNs showed more effective uptake with respect to the naked by tumor cells that overexpress integrin receptors (but not for non-overexpressing ones). In contrast, at higher concentrations, the non-specific cell membrane protein–particle interactions are promoted and coupled to specific and target mediated uptake. Visual observations by fluorescence microscopy suggested that SFNs bind to integrin receptors on the cell surface and are then internalized by endocytosis. Overall, SFN functionalization provided in vitro active targeting for site-specific delivery of anticancer drugs, boosting activity and sparing healthy organs. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Multifunctional nanomaterials integrating therapeutic and imaging modalities in one platform have opened a new era in the present therapeutic strategies. In the present study, a multifunctional silk fibroin-based carrier has been designed for the delivery of antioxidant and imaging agents. One-step desolvation method was used to prepare sulforaphane (antioxidant drug) loaded silk fibroin nanoparticles (SFSNPs). These anionic SFSNPs were further coupled with cationic cerium oxide nanoparticles (CeNPs) and PEI passivated carbon dots (CDs) to form self-assembled CeNP-CD@SFSNPs nanocomposites. CDs were synthesized from mulberry leaves (Morus indica) as green source of carbon and bPEI as a passivating agent to get positively charged CDs. The CDs functioned as molecular probes by emitting green fluorescence while the presence of CeNPs augmented the antioxidant potential due to their unique redox property. Time-dependent in vitro release of sulforaphane was fast in acidic pH than under normal physiological conditions. Cytotoxicity studies were performed on L132 normal epithelial lung cell lines and A549 lung cancer cell lines to analyze the toxicity of the nanocomposites. Green fluorescence from the CDs facilitated in fluorescence microscopic imaging and cellular uptake studies. ROS scavenging capability was analyzed by exposing cells to H 2 O 2 stress using flow cytometry and DCFH-DA staining. Overall, the synthesized CeNP-CD@SFSNPs nanocomposites efficiently reduced ROS levels by simultaneously enabling imaging of the cells. Thus, this CeNP-CD@SFSNPs nanocomposite could be a potential candidate for simultaneous imaging and drug delivery against oxidative stress. Image 1 • Single step hydrothermal synthesis of carbon dots (CDs) from mulberry leaves. • Cerium oxide nanoparticles and CDs coupled to sulforaphane loaded silk fibroin nanoparticles. • Imaging and antioxidant activity of theranostic nanocomposites was validated by various assays. [ABSTRACT FROM AUTHOR]

Objective: the objective of the present work is to study the effectiveness of treatment with silk fibroin nanoparticles loaded with resveratrol in experimental periodontitis in a diabetic rat model. Introduction: Periodontitis is an inflammatory pathology highly related to other diseases, such as type II diabetes. Both diseases have a specific inflammatory condition, with Interleukin (IL)-6, IL-1β and Transforming Grow Factor (TGF)-1β being the most relevant proinflammatory factors. Silk fibroin (SF) nanoparticles loaded with resveratrol (Res-SFN) are a new alternative as a treatment. Methods: 40 diabetic Sprague Dawley male rats were used and periodontitis was induced by ligation. The animals were divided into 5 treatment groups, and 1 mL of treatment was administered once a day for 4 weeks. The groups were: I: Carboxymethyl cellulose (CMC) 0.8%, II: CMC 0.8% + SF 1%, III: CMC 0.8% + RES-SFN 3 mg/mL, IV: CMC 0.8% + SF 1% + RES-SFN 3 mg/mL, V: Water. A peripheral blood sample was taken every week to quantify the inflammatory profile by ELISA (IL-6, IL-1β and TGF-1β). After 4 weeks the sacrifice was carried out and biopsies of the gum were taken. Results: Treatment with SF and RES-SFN reduced the amount of chemical inflammation mediators (with the exception of IL-1β in comparisons I-IV and II-IV (p > 0.05)), as well as the anatomopathological variables linked to it, in a significant way (p < 0.05). Conclusion: treatment with RES-SFN has reduced local inflammation in this experimental periodontitis model. [ABSTRACT FROM AUTHOR]

In this study, the silk fibroin nanoparticle (SFNP) was used as a nanoadjuvant in combination with a multiepitope-based vaccine for urinary tract infection (UTI). Nanoparticles containing the fusion protein were analyzed for physicochemical properties, toxicity, release profile, and in vivo potency. The synthesized nanovaccine showed a spherical shape with a mean particle size of 180 nm and an encapsulation efficiency of 88%. Antigen release from SFNPs was 18% after 42 days. The SFNPs showed a zeta potential of -29 mV and had no toxic effect on the L929 cells in vitro. SFNPs in the vaccine formulations promoted humoral and cellular (IFN-γ, IL-4, and IL-17) immune responses in comparison to controls. Immunization of mice with antigen-encapsulated SFNPs significantly increased the total IgG as well as IgG2a/IgG1 ratio. In addition, this formulation triggered concurrently type 1 (Th1) and type 2 (Th2) immune responses, with a Th1-polarized response. Furthermore, highly effective protection of the bladder and kidney against experimental UTI was obtained by using the nanoadjuvant containing the antigen for 6 months. The results demonstrated that SFNPs can be proposed as potent adjuvants or vaccine carriers to develop new and more effective nanovaccine formulations in the future.

This work aims to provide an effective and novel dual tool for the biodistribution studies of biopolimeric nanoparticles by using modified silk fibroin nanoparticles as a model. This is an indispensable step in the evaluation of the applicability of biopolymeric nanoparticles as drug delivery systems. In this work, we report a new facile method for radiolabeling silk fibroin nanoparticles conjugated to the chelating agent diethylenetriamine pentaacetic acid and tagged with fluorescein isothiocyanate. Nanoparticles were characterized by means of dynamic light scattering, scanning electron microscopy, and infrared and fluorescence spectroscopy. The in vitro studies included stability in biological media and evaluation of the cytotoxicity of the nanoparticles in a cell culture. The in vivo study was focused on a scintigraphic study over 24 h conducted on New Zealand rabbits, after intra-articular injection of [ 111 In]In-nanoparticles containing 8.03 ± 0.42 MBq. Biodistribution of the nanoparticles was also assessed ex vivo by fluorescence microscopy of post mortem biopsied organs. This radiolabeling method was reproducible and robust with high radiolabeling efficiency (∼80%) and high specific activity suitable for in vivo studies. Radiolabeled nanoparticles, having a hydrodynamic radius of 113.2 ± 2.3 nm, a polydispersity index of 0.101 ± 0.015, and a Z -potential of -30.1 ± 2.0 mV, showed an optimum retention in the articular space, without activity clearance up to 24 h post injection. Thus, an easy and robust radiolabeling method has been developed, and its applicability is demonstrated in vitro and in vivo studies, showing its value for future investigation of silk fibroin nanoparticles as versatile and stable (steady) local drug delivery systems for consideration as a therapeutic option, particularly in the treatment of joint disorders.

Silk fibroin (SF) scaffolds are widely used in tissue engineering due to their biocompatibility and slow biodegradability. However, the relatively stiff mechanical properties and low permeability of these systems can limit some applications. In this study, a new type of water-stable silk sponge (ASF-PEG-S) was obtained by inducing nanoparticle (50-300 nm in diameter) formation in SF solution by autoclaving followed by freeze-drying and rinsing the dry sponges with low-molecular weight (400 Da) polyethylene glycol (PEG400) to induce SF β-sheet structure formation and thus stability in water. With further extraction, the SF nanoparticles embedded in the sponges were removed, leaving nanopores in the walls of round-shaped micro-size pores. The unique pore structure resulted in enhanced permeability and flexibility of ASF-PEG-S when compared to other types of SF sponges, especially with respect to commonly used methanol-annealed SF sponges. In addition, ASF-PEG-S absorbed water nearly 40 times more than its dry weight, while the methanol-annealed sponges absorbed half this amount. When human fibroblasts were seeded and cultured on ASF-PEG-S versus traditional SF methanol-processed sponges, improved cell encapsulation, distribution, and consistency in growth were observed, suggesting utility in tissue engineering and tissue repair applications in the future.

Several studies have stated that the process used for sericin removal, or degumming, from silk cocoons has a strong impact in the silk fibroin integrity and consequently in their mechanical or biochemical properties after processing it into several biomaterials (e.g. fibers, films or scaffolds) but still, there is a lack of information of the impact on the features of silk nanoparticles. In this work, silk cocoons were degummed following four standard methods: autoclaving, short alkaline (Na2CO3) boiling, long alkaline (Na2CO3) boiling and ultrasounds. The resultant silk fibroin fibers were dissolved in the ionic liquid 1-ethyl-3-methylimidazolium acetate and used for nanoparticle synthesis by rapid desolvation in polar organic solvents. The relative efficiencies of the degumming processes and the integrity of the resulting fibroin fibers obtained were analyzed by mass loss, optical microscopy, thermogravimetric analysis, infrared spectroscopy and SDS-PAGE. Particle sizes and morphology were analyzed by Dynamic Light Scattering and Field Emission Scanning Electronic Microscopy. The results showed that the different treatments had a remarkable impact on the integrity of the silk fibroin chains, as confirmed by gel electrophoresis, which can be correlated with particle mean size and size distribution changes. The smallest nanoparticles (156 ± 3 nm) and the most negative Z potential (−30.2 ± 1.8 mV) were obtained with the combination of long treatment (2 h) of boiling in alkaline solution (Na2CO3 0.02 eq/L). The study confirms that parameters of the process, such as composition of the solution and time of the degumming step, must be controlled in order to reach an optimum reproducibility of the nanoparticle production. [ABSTRACT FROM AUTHOR]

Celastrol (CL), a bioactive compound isolated from Tripterygium wilfordii, has demonstrated bioactivities against a variety of diseases including cancer and obesity. However, its poor water solubility and rapid in vivo clearance limit its clinical applications. To overcome these limitations, nanotechnology has been employed to improve its pharmacokinetic properties. Nanoparticles made of biological materials offer minimal adverse effects while maintaining the efficacy of encapsulated therapeutics. Silk fibroin (SF) solution was prepared successfully by extraction from the cocoons of silkworms, and a final concentration of 2 mg/mL SF solution was used for the preparation of CL-loaded SF nanoparticles (CL-SFNP) by the desolvation method. A stirring speed of 750 rpm and storage time of 20 h at −20 °C resulted in optimized product yield. A high-performance liquid chromatography (HPLC) method was developed and validated for the analysis of CL in rat plasma in terms of selectivity, linearity, intra-/inter-day precision and accuracy, and recovery. No interference was observed in rat plasma. Linearity in the concentration range of 0.05–5 µg/mL was observed with R2 of 0.999. Precision and accuracy values were below the limit of acceptance criteria, i.e., 15% for quality control (QC) samples and 20% for lower limit of quantification (LLOQ) samples. Rats were given intravenous (IV) administration of 1 mg/kg of pure CL in PEG 300 solution or CL-SFNP. The pharmacokinetic profile was improved with CL-SFNP compared to pure CL. Pure CL resulted in a maximum concentration (Cmax) value of 0.17 µg mL−1 at 5 min following administration, whereas that for CL-SFNP was 0.87 µg mL−1 and the extrapolated initial concentrations (C0) were 0.25 and 1.09 µg mL−1, respectively, for pure CL and CL-SFNP. A 2.4-fold increase in total area under the curve (AUC0-inf) (µg h mL−1) was observed with CL-SFNP when compared with pure CL. CL-SFNP demonstrated longer mean residence time (MRT; 0.67 h) than pure CL (0.26 h). In conclusion, the preparation of CL-SFNP was optimized and the formulation demonstrated improved pharmacokinetic properties compared to CL in solution following IV administration. [ABSTRACT FROM AUTHOR]