Abstract For over a century, artificial silk spinning has pursued the exceptional mechanical performance of natural fibres, largely through a molecular-centric strategy. Limitations in artificial systems have been attributed to insufficient molecular weight, incomplete sequence architecture, or loss of native terminal domains, factors also thought to disrupt liquid-liquid phase separation (LLPS) and hierarchical assembly. LLPS is proposed to concentrate and pre-organise silk proteins, facilitating alignment during spinning and formation of hierarchical structures that underpin mechanical performance. Here, we reevaluate this paradigm through comparisons of regenerated silk fibroin (RSF), recombinant silk proteins, and regenerated undegummed silk (RUS). Advances across these systems have substantially narrowed the molecular gap with native silk, and under optimised conditions, all can produce fibres with comparable mechanical properties. While LLPS and hierarchical organisation can be induced in RSF and recombinant systems, these features do not consistently improve mechanics, suggesting native-like assembly alone is insufficient. RUS, which retains multicomponent interactions, most closely reflects the native system, yet still exhibits distinct rheology. Collectively, this indicates that artificial dopes lack compositional complexity and ability to respond to dynamic physiochemical gradients of the silk gland. Future progress in artificial spinning will require reconstructing the multicomponent, non-equilibrium environment governing silk assembly in nature.

Silk sericin, a hydrophilic protein traditionally discarded during silk degumming, holds significant potential for biomedical applications due to its antioxidant, antibacterial, and biocompatible properties. Conventional chemical degumming produces a high yield but degrades sericin, impacting its properties. Other extraction methods utilizing denaturants such as guanidine hydrochloride produce high-quality sericin but at an extremely low yield. This study introduces an innovative mechanical extraction method that combines cutter milling with shear homogenization in water to isolate sericin from cocoons without the use of chemicals. Through careful control of shear intensity and thermal management, the process achieved a yield of 7.0 +/- 1.4%, nearly 3-fold higher than guanidine hydrochloride extraction. Sodium dodecyl-sulfate polyacrylamide gel electrophoresis of the extracted sericin revealed minimal degradation, with distinct bands ranging from similar to 20 kDa to over 200 kDa. Unlike alkaline-degummed sericin, the extract produced a thick hydrogel. Proteomic analysis of the extract revealed a unique composition, with a high proportion of Sericin 3, whereas chemical extraction yielded more Sericin 1. The ability to selectively extract sericin fractions may deepen understanding of their functional roles and enable targeted strategies to precisely tune hydrogel properties.

Liquid-liquid phase separation (LLPS) is a fundamental principle in protein assembly, directing processes ranging from the formation of membraneless organelles in biological cells to the self-assembly of silk proteins. Silk LLPS is typically studied in phosphate buffers, but it can also occur spontaneously without solvent modification. By directly comparing these two pathways, we demonstrate that phosphate ions trigger assemblies fundamentally distinct from natural, salt-free LLPS. The natural process, driven by silk's heterogeneous and amphiphilic character, yields dynamic, easily disrupted condensates. In contrast, phosphate ions promote interactions that generate larger, denser, and far more stable condensates that persist after drying. We also show that fibroin distributes throughout the outer phase in natural LLPS but migrates exclusively to the dense phase with phosphate ions. This establishes that phosphate ions actively reshape both the composition and properties of silk condensates and should be wielded with care. Our insights provide critical clues into how metal ions guide silk solidification in vivo and will inform the design of improved regenerated silk feedstocks for spinning. More broadly, this work advances our understanding of protein self-assembly, with potential implications for counteracting pathological protein aggregation in disease.

The self-assembly of β-sheet rich proteins such as silk are triggered by physiochemical factors such as salts, pH, and flow. Anions, in particular, have been shown to significantly influence protein self-assembly in amyloids and spider silk; however, their effect on silkworm silk has not yet been investigated. This study evaluated the influence of metal ions on silk gelation kinetics by utilizing a recently developed system to dissolve undegummed Bombyx mori cocoons. While the influence of cations agreed with previous observations, with divalent cations delaying gelation more than monovalent ions, we report that anions are more influential, with phosphate and sulfate anions accelerating gelation, regardless of the valency of the associated cation, while bromide was found to delay it. Interestingly we discovered chloride stabilized silk regardless of the cation. Using these results, a linear relationship was established between gelation time and apparent dynamic hydration number, a quantitative measure of salt-water interactions. This relationship more accurately predicted silk behavior than the traditionally used Hofmeister series. Thus, it was possible to accurately predict gelation behavior for any salt for which this number has been calculated, allowing for better designs of systems that require fine control of gelation such as wet-spinning and 3D printing. This work demonstrates the significance of anions in controlling silkworm silk stability, expanding upon the currently perceived mechanism for self-assembly.

Microfibrillated silk (MFS) is an emerging class of silk materials produced by directly exfoliating silk fibres, offering a top-down alternative to traditional regenerated silk processing methods. They have the potential to be used as biomaterials, particularly in tissue engineering and regenerative medicine. This study used a unique tuneable top-down approach to produce different MFS suspensions. These were then fabricated into protein papers using two scalable methods, casting and vacuum filtration, to examine how processing and fabrication together influenced the final material properties and associated cellular responses. MFS suspensions were prepared under three processing levels using mechanical processing alone or combined with acid pre-treatment with each level yielding increasingly finer fibrils. The level of fibrillation significantly affected fibre morphology and mechanical strength, while the fabrication method mainly influenced surface roughness and bilayer characteristics. Papers made with mechanical processing alone had the highest strength. Cast papers revealed a surface difference, with a rough top surface and smooth bottom surface, while vacuum-filtered papers had uniform roughness on both sides. These surface differences, along with the degree of fibrillation, impacted cell attachment and organization, with cast papers from acid-pretreated and shear-homogenised MFS showing the best biological outcomes. Controlling MFS processing and assembly techniques to form papers enables the design of silk-based materials tailored for various biomedical applications.

This study demonstrates the use of spray drying as a versatile processing technique to produce regenerated silk powders with a controllable particle size and solubility. After overcoming silk's shear sensitivity and establishing a usable processing window, semicrystalline silk powders were produced. The impact of silk properties and spray drying conditions on powder properties was then explored. Spray drying produced spherical hollow or collapsed particles similar to other spray dried proteins; particle size could be controlled from a d(50) of less than 5 to almost 40 μm through altering feedstock concentration. Interestingly, the silk secondary structure, which typically dictates silk solubility, was resilient to changes in spray drying conditions: all powder samples ranged from 39 to 42% β-sheet content. Yet despite these findings, silk solubility could be controlled from less than 4% to nearly 60% purely by changing fibroin molecular weight and, to a lesser extent, concentration. Given the commercial viability of spray drying, this study demonstrates significant potential for the production of powders with controllable properties for a range of possible applications, from biomaterials to food or cosmetic applications.

Degradable hemostatic materials such as gelatin foam are widely used as middle ear packing materials to support tympanic membrane grafts; while safe and biocompatible, such materials have limited mechanical stability when wet. Here, we demonstrate the fabrication of silk-silk composite foams by mixing microfibrillated silk with a low molecular weight silk solution glue to produce water-stable and highly porous foams that retain their structure when wet. The foams exhibited excellent (over 2000%) water absorption, more than 5 times higher than Gelfoam, a commercial hemostatic packing material, and hence higher loading of antibiotics such as ciprofloxacin. Moreover, unlike Gelfoam, ciprofloxacin could be encapsulated into the composite foams during fabrication to produce a dried, off-the-shelf, drug-eluting packing material. This enabled sustained drug release for over 16 days. The addition of microfibers significantly improved shape retention after annealing: samples containing at least a 1:1 mix of silk solution to microfibers showed no shrinkage after annealing, while control foams made from just silk solution shrank by at least 25%. These hybrid foams have immense potential for use as dual-function factor-concentrating hemostatic materials with drug delivery capacity for middle ear packing.

The remarkable toughness (>70 MJ m-3) of silkworm silk is largely attributed to its hierarchically arranged nanofibrillar nanostructure. Recreating such tough fibers through artificial spinning is often challenging, in part because degummed, dissolved silk is drastically different to the unspun native feedstock found in the spinning gland. The present work demonstrates a method to dissolve silk without degumming to produce a solution containing undegraded fibroin and sericin. This solution exhibits liquid-liquid phase separation above 10% (wt/wt), a behavior observed in the silk gland but not in degummed silk solutions to date. This partitioning enhances the stability of the undegummed solution, delaying gelation two-fold compared with degummed silk at the same concentration. When spun under identical conditions, undegummed solutions produces fibers 8× stronger and 218× tougher than degummed silk feedstocks. Through ultrasonication, undegummed wet spun fibers are seen to possess hierarchical structure of densely packed ≈20 nm nanofibrils, similar to native silks, although completely absent from fibers wet-spun from degummed silk solutions. This work demonstrates that the preservation of molecular weight, presence of sericin and stimulation of liquid-liquid phase separation underpin a new pathway to recreate a hierarchical fiber with structures akin to native silk.

Biorefinery, the process of converting biomass to value-added materials, is gaining significant interest because of the depletion of natural resources and an increasing awareness of the need for sustainable development and material production. Silk sericin, an abundantly available byproduct of the sericulture industry, has been identified for biorefinery usage since it is biodegradable, bioavailable, pH and temperature-responsive, naturally photoluminescent, has antioxidant properties, and is a GRAS and FDA-safe food additive. This Perspective examines the properties of sericin and its sources and discusses currently explored applications, including as a delivery system for drugs or genetic material, for wound healing, tissue engineering, media supplements, cosmetology, packaging materials, edible coating, and as bio adhesives. This Perspective examines the sources of sericin, its properties, and some of the currently explored applications. These include biomedical applications such as a delivery system for drugs or genetic material, wound healing, tissue engineering, media supplements, cosmetology, packaging materials, edible coating, and as bio adhesives. Finally, we highlight the recent developments in sericin modification to functionalize it for specific applications; this is a promising future direction that will further enhance the potential use of this valuable biomass.

Cardiovascular diseases are a major global health challenge. Blood vessel disease and dysfunction are major contributors to this healthcare burden, and the development of tissue-engineered vascular grafts (TEVGs) is required, particularly for the replacement of small-diameter vessels. Silk fibroin (SF) is a widely used biomaterial for TEVG fabrication due to its high strength and biocompatibility. However, the stiffness of SF is much higher than that of native blood vessels (NBVs), which limits its application for vascular tissue engineering. In this study, SF was plasticized with glycerol to produce TEVGs exhibiting similar stiffness and ultimate tensile strength to those of NBVs. The electrospun SF/glycerol TEVGs exhibited mechanical properties comparable to NBVs and supported the in vitro proliferation of essential vascular cells-endothelial and smooth muscle cells. After 5 days of culture, the TEVGs exhibited an endothelial monolayer in the lumen, demonstrating their potential for functional vascular tissue regeneration. Our study demonstrates the feasibility of producing TEVGs from SF with tailored mechanical properties, paving the way for more functional and durable TEVGs for future clinical applications.

Microfibrillated silk (MFS) is an emerging class of silk-based materials with diverse properties and potential uses. While regenerated silk materials produced using the “bottom-up” approach require complete silk dissolution, the “top-down” microfibrillation of silk fibers only partially disrupts silk’s hierarchical structure to release fibrils that largely retain native silk’s nanocrystalline structure. We have used here the term MFS for exfoliated fibrils with an average diameter of more than 100 nm and the term nanofibrillated silk (NFS) for finer fibrils with a diameter of less than 100 nm. However, these terms microfibrillation, nanofibrillation, MFS, NFS, or fibrillated silk are used interchangeably in the published literature and overall represent exfoliated silk materials. This chapter provides an overview of the processing required to exfoliate silk. Since their use in tissue engineering and biomedicine is in its infancy, this chapter also discusses established bottom-up nanofibrous materials that have been developed for biomedical applications since MFS/NFS are expected to be used in many such applications. Finally, given the significant differences in properties, we provide our perspective on the potential usefulness of MFS in numerous applications including next-generation biomedical products.

The development of stimuli-responsive nano-formulation involves the reduction of losses caused by commercial fertilizers. It can provide controlled release kinetics that enhances the permeability, stability, and solubility, thus further improving the efficiency of nano-formulation by extending the duration of premature degradation of active ingredients under harsh conditions. These formulations are designed to effectively deliver significant amounts of active ingredients in response to abiotic stresses that act as inducers, which led to targeted and controlled release mechanisms. This chapter discusses the status of the stimuli-responsive systems with the prospect to be used in agriculture, highlighting the types of stimuli-responsive systems and their functionality in the release of active ingredients. The future trends involving the stimuli-responsive nano-formulations used in agriculture need to be explored extensively. Thus proper guidelines should be set up for the field application of stimuli-responsive nano-formulations with minimum impact on the environment.

In recent years, significant progress has been made in both three-dimensional (3D) printing technologies and the exploration of silk as an ink to produce biocompatible constructs. Combined with the unlimited design potential of 3D printing, silk can be processed into a broad range of functional materials and devices for various biomedical applications. The ability of silk to be processed into various materials, including solutions, hydrogels, particles, microspheres, and fibers, makes it an excellent candidate for adaptation to different 3D printing techniques. This review presents a didactic overview of the 3D printing of silk-based materials, major categories of printing techniques, and their prototyping mechanisms and structural features. In addition, we provide a roadmap for researchers aiming to incorporate silk printing into their own work by summarizing promising strategies from both technical and material aspects, to relate state-of-the-art silk-based material processing with fast-developing 3D printing technologies. Thus, our focus is on elucidating the techniques and strategies that advance the development of precise assembly strategies for silk-based materials. Precise printing (including high printing resolution, complex structure realization, and printing fidelity) is a prerequisite for the digital design capability of 3D printing technology and would definitely broaden the application era of silk, such as complex biomimetic tissue structures, vasculatures, and transdermal microneedles.

Hybrid protein-copper nanoflowers have emerged as promising materials with diverse applications in biocatalysis, biosensing, and bioremediation. Sericin, a waste biopolymer from the textile industry, has shown potential for fabricating such nanoflowers. However, the influence of the molecular weight of sericin on nanoflower morphology and peroxidase-like activity remains unexplored. This work focused on the self-assembly of nanoflowers using high- and low-molecular-weight (HMW and LMW) silk sericin combined with copper(II) as an inorganic moiety. The peroxidase-like activity of the resulting nanoflowers was evaluated using 2,2 '-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and hydrogen peroxide (H2O2). The findings revealed that high-molecular-weight sericin hybrid nanoflowers (HMW-ShNFs) exhibited significantly higher peroxidase-like activity than low-molecular-weight sericin hybrid nanoflowers (LMW-ShNFs). Furthermore, HMW-ShNFs demonstrated superior reusability and storage stability, thereby enhancing their potential for practical use. This study also explored the application of HMW-ShNF for ciprofloxacin degradation to address the environmental and health hazards posed by this antibiotic in water. The results indicated that HMW-ShNFs facilitated the degradation of ciprofloxacin, achieving a maximum degradation of 33.2 +/- 1% at pH 8 and 35 degrees C after 72 h. Overall, the enhanced peroxidase-like activity and successful application in ciprofloxacin degradation underscore the potential of HMW-ShNFs for a sustainable and ecofriendly remediation process. These findings open avenues for the further exploration and utilization of hybrid nanoflowers in various environmental applications.

IntroductionFertilizer management is crucial to maintaining a balance between environmental health, plant health, and total crop yield. Farmers are overutilizing fertilizers with a mind set to enhance the productive capacity of the field, which adversely impacts soil fertility and causes serious environmental hazards. To mitigate the issues of over-utilization of fertilizers, controlled-release fertilizers were developed using nitrogen fertilizer (ammonium chloride) loaded on cellulose nanofibres (named CNF*N).MethodologyIn this study, the effects of CNF*N were compared with commercial nitrogen fertilizer (ammonium chloride) on Vigna radiata (Mung) under greenhouse conditions. The pot experiment was conducted using six treatments: first treatment was control, where the plant was cultivated (T1); second treatment was T2, where the plant was cultivated with CNF to determine the impact of CNF on the plant; third was T3 where commercial ammonium chloride (24 mg/ 2 kg soil) was added to the plant; fourth was T4, where the plant was loaded with CNF, viz. CNF*N contains 4.8 mg of nitrogen; fifth was T5 CNF*N pellet contains 12 mg of nitrogen, and the last sixth treatment (T6) where CNF*N pellet containing 24 mg of nitrogen.ResultsIt indicated that the growth parameters were best achieved in T6 treatment. Plant height was at its maximum in the T6 treatment (44.4 ±0.1cm) after the second harvest, whereas the minimum plant height was observed in T1, which was 39.1 ±0.1 cm. Root-to-shoot weight ratio was also maximum in T6 (0.183± 0.02) and minimum in T1 (0.07± 0.01) after second harvesting. The significant difference among the treatments was determined with Tukey’s honestly significant difference (HSD). The nitrogen content (available and total) was significantly higher in the T4, T5, and T6 treatments (0.22, 0.25, and 0.28%) as compared to the control treatments (T1 (0.12%), T2 (0.13%), and T3 (0.14%) during the second harvesting stage (90 days), as nitrogen plays a crucial role in the development of vegetative growth in Vigna radiata. The rate of controlled-release nitrogen-fertilizer was found to be optimal in terms of plant growth and soil nutrients; hence, it could potentially play a crucial role in improving soil health and the yield of the crop.

Biopolymers play a critical role as scaffolds used in tendon and ligament (TL) regeneration. Although advanced biopolymer materials have been proposed with optimised mechanical properties, biocompatibility, degradation, and processability, it is still challenging to find the right balance between these properties. Here, we aim to develop novel hybrid biocomposites based on poly(p-dioxanone) (PDO), poly(lactide-co-caprolactone) (LCL) and silk to produce high-performance grafts suitable for TL tissue repair. Biocomposites containing 1-15% of silk were studied through a range of characterisation techniques. We then explored biocompatibility through in vitro and in vivo studies using a mouse model. We found that adding up to 5% silk increases the tensile properties, degradation rate and miscibility between PDO and LCL phases without agglomeration of silk inside the composites. Furthermore, addition of silk increases surface roughness and hydrophilicity. In vitro experiments show that the silk improved attachment of tendon-derived stem cells and proliferation over 72 h, while in vivo studies indicate that the silk can reduce the expression of pro-inflammatory cytokines after six weeks of implantation. Finally, we selected a promising biocomposite and created a prototype TL graft based on extruded fibres. We found that the tensile properties of both individual fibres and braided grafts could be suitable for anterior cruciate ligament (ACL) repair applications.

Currently, the packaging material is composed of fossil fuel-based plastics, which are non-biodegradable. Therefore, the biodegradable coating material developed from cellulose can be explored for applications like packaging, cutlery, bowls, straws, etc. The aim of the present study was to develop an agro-waste-based sustainable coating material using a waste-to-wealth approach. The study focused on transforming cellulose and cellulose nanofibre (CNF) derived from rice straw into biodegradable polymeric coatings. The cellulose and CNF were extracted from the rice straw using the alkali method and blended with gelatin, agar, and varied glycerol concentrations to initiate the study. Four polymeric coating variants were developed. Thus, the coating named NCG-2.5, incorporated with 1.2 g CNF, 2.5 ml glycerol, 1.15 g gelatin, and 0.55 g agar, exhibited significant improvements in properties as compared to the other samples. The polymeric coatings were characterized for their surface morphology, roughness, solubility, and physico-chemical properties using FTIR spectroscopy and TGA analysis. The NCG-2.5 exhibited the best potential in terms of promising coating and biodegradability properties, with 97

Silk from silkworms and spiders is an exceptionally important natural material, inspiring a range of new products and applications due to its high strength, elasticity, and toughness at low density, as well as its unique conductive and optical properties. Transgenic and recombinant technologies offer great promise for the scaled-up production of new silkworm- and spider-silk-inspired fibres. However, despite considerable effort, producing an artificial silk that recaptures the physico-chemical properties of naturally spun silk has thus far proven elusive. The mechanical, biochemical, and other properties of pre-and post-development fibres accordingly should be determined across scales and structural hierarchies whenever feasible. We have herein reviewed and made recommendations on some of those practices for measuring the bulk fibre properties; skin-core structures; and the primary, secondary, and tertiary structures of silk proteins and the properties of dopes and their proteins. We thereupon examine emerging methodologies and make assessments on how they might be utilized to realize the goal of developing high quality bio-inspired fibres.