Lignin nanoparticles formation by multiscale structure control to regulate morphology and their adsorption, nucleation, and growth on chitin nanofibers.

The lignin nanoparticles (LNPs) synthesis relies on lignin polymers with heterogeneous molecules and properties, which impose significant limitations on the preparation and property regulation. The multiscale structure of lignin from monomers to oligomers, provides a potential pathway for precise regulation of its physical and chemical properties. The study addresses this challenge by employing coniferyl alcohol and sinapyl alcohol as monomers and separately utilizing the Zulaufverfaren (ZL) and Zutropfverfaren (ZT) methods to synthesize different types of lignin dehydrogenation polymers (DHPs) including guaiacyl (G)-ZL-DHP, G-ZT-DHP, syringyl (S)-ZL-DHP, and S-ZT-DHP. The investigation highlights the chemical bonds as essential components of lignin primary structure. Additionally, the secondary structure is influenced by branched and linear molecular structures. G unit provides some branching points, which are utilized and amplified in the ZL process of DHPs synthesis. The branched DHPs aggregate at the edge and form rod-like LNPs. While linear DHPs aggregate around the center, presenting polygonal LNPs. The study identifies that the branched LNPs, characterized by more surface charges and lower steric hindrance, can form a stable complex with chitin nanofibers. Emulsions with varying oil-to-water ratios were subsequently prepared, opening a new window for the application of LNPs in fields such as food and cosmetics.

Role of chitin synthases CHS1 and CHS2 in biosynthesis of the cyst wall of Cryptocaryon irritans.

Cryptocaryon irritans, a protozoan parasite that infects marine fish, is characterized by a complex life cycle that includes a cyst-forming reproductive phase. However, the composition of the cyst wall and mechanism of its formation remain unclear. In this study, we identified chitin as a key component of the cyst wall using calcofluor white and wheat germ agglutinin, with Fourier-transform infrared spectroscopy confirming its ß-form structure. Two chitin synthase genes, CHS1 and CHS2, were identified as being expressed throughout the life cycle and show close phylogenetic relationships with chitin synthase from ciliates. Incubation with specific anti-CHS1 and -CHS2 antibodies significantly reduced both the thickness and chitin content of the cyst wall, highlighting the critical role of these enzymes in chitin biosynthesis. Treatment with benzoylureas, which inhibit chitin synthesis, caused thinning of the cyst wall and downregulation of CHS gene expression, resulting in an 84 % reduction in the hatching rate after treatment with 0.01 mM CuSO4 compared with control tomonts. Western blot analysis demonstrated that recombinant CHS proteins are immunogenic, and tomonts from CHS-immunized grouper exhibited reduced size. These findings bridge a crucial knowledge gap in understanding of the C. irritans cyst wall and highlight promising targets for infection prevention and control strategies.

Differences in transcriptomic responses upon Phytophthora palmivora infection among cultivars reveal potential underlying resistant mechanisms in durian.

BACKGROUND: Phytophthora palmivora is a devastating oomycete pathogen in durian, one of the most economically important crops in Southeast Asia. The use of fungicides in Phytophthora management may not be a long-term solution because of emerging chemical resistance issues. It is crucial to develop Phytophthora-resistant durian cultivars, and information regarding the underlying resistance mechanisms is valuable for smart breeding programs. RESULTS: In this study, we conducted RNA sequencing (RNA-seq) to investigate early gene expression responses (at 8, 24, and 48 h) after the P. palmivora infection in three durian cultivars, which included one resistant cultivar (Puangmanee; PM) and two susceptible cultivars (Monthong; MT and Kradumthong; KD). We performed co-expression and differential gene expression analyses to capture gene expression patterns and identify the differentially expressed genes. The results showed that genes encoding heat shock proteins (HSPs) were upregulated in all infected durians. The expression levels of genes encoding HSPs, such as ERdj3B, were high only in infected PM. A higher level of P. palmivora resistance in PM appeared to be associated with higher expression levels of various genes encoding defense and chitin response proteins, such as lysM domain receptor-like kinases. MT had a lower resistance level than PM, although it possessed more upregulated genes during P. palmivora infection. Many photosynthetic and defense genes were upregulated in the infected MT, although their expression levels were lower than those in the infected PM. KD, the least resistant cultivar, showed downregulation of genes involved in cell wall organization or biogenesis during P. palmivora infection. CONCLUSIONS: Our results showed that the three durian cultivars exhibited significantly different gene expression patterns in response to P. palmivora infection. The upregulation of genes encoding HSPs was common in all studied durians. The high expression of genes encoding chitin response proteins likely contributed to P. palmivora resistance in durians. Durian susceptibility was associated with low basal expression of defense genes and downregulation of several cell wall-related genes. These findings enhance our understanding of durian resistance to Phytophthora infection and could be useful for the development of elite durian cultivars.

Hydrothermal preparation of pharmaceuticals adsorbents from chitin and chitosan: Optimization and mechanism.

Hydrothermal treatment of fishery waste-derived chitin (CT) and chitosan (CS) was performed to prepare biochar adsorbents for the removal of pharmaceuticals of environmental concern. By systematically studying the effect of treatment conditions on the biochar structure, the correlation between biochar properties and the adsorption capacities was clarified to optimize the adsorption performance. CS hydrochars obtained by lower-temperature treatment showed high adsorption capacities for the pharmaceuticals having carboxyl groups attributed to the electrostatic binding. A decrease in the density of available amines in CS hydrochars prepared at higher temperatures resulted in lower adsorption capacities and the manifestation of different adsorption mechanisms based on hydrophobic and π-π interactions. CT hydrochars showed lower adsorption capacities than CS hydrochars due to inefficient carbonization and lack of adsorption sites. The hydrochar adsorbents prepared in this study address simultaneously the problems of marine waste bioresource utilization and environmental cleaning from the emergent pollutants.

Water-in-water emulsions stabilized by nano-chitin.

A water-in-water (W/W) emulsion consists of microdroplets was formed by the spontaneous liquid-liquid separation by mixing polyacrylic acid and chitosan oligosaccharide in water, and these microdropletes were stabilized by nano-chitin, formed water-in-water Pickering emulsions. By taking the advantage of interfacial adsorption of nano-chitin, the W/W emulsion droplets composed of polyacrylic acid/chitosan oligosaccharide (COS/PAA) polyelectrolyte coacervate were successfully stabilized. Research results indicated that composite microspheres were formed by the nano-chitin stabilized COS/PAA emulsion, and the size of these composite microspheres was related to the concentration and morphology of the nano-chitin. As the concentration of nano-chitin increases, the size of the composite microspheres first increases and then decreases, gradually becoming more uniform; whereas a decrease in the length of nano-chitin will lead to an increase in the size of the composite microspheres. The formation of composite microspheres may be due to the electrostatic interactions between nano-chitin and the emulsion droplets. In addition, the composite microspheres exhibit the best release effect for berberine hydrochloride at a pH value of 3.

Chitinase and Deacetylase-Based Chitin-Degrading Bacteria: One-Pot Cascade Bioconversion of Chitin to Chitooligosaccharides.

Cascade conversion of chitin into soluble and functional chitooligosaccharides has gained great attention. However, the biotransformation route is still limited to the low catalytic performances of chitin deacetylases (CDAs) and complicated procedures. In this study, a CDA from Arthrobacter sp. Jub115 (ArCDA) was identified and characterized, which showed a higher catalytic stability than the reported CDAs, with residual activity of 80.49%, 71.12%, and 56.09% after incubation at 30, 35, and 40 °C for 24 h, respectively. Additionally, ArCDA was identified to have a broad substrate spectrum toward ß-chitin and N-acetyl chitooligosaccharides. Moreover, an engineered chitin-degrading bacteria (CDB) with cell-surface-displayed deacetylase ArCDA and chitinase SaChiB was constructed to simplify catalysis procedures, facilitating the chitobiose production of 294.30 ± 16.43 mg/L in 10 h. This study not only identified a CDA with the desirable catalytic performance but also provided a strategy for constructing CDB, facilitating the high-value utilization of chitin.

A comprehensive review of chitosan-based functional materials: From history to specific applications.

Chitosan (CTS), a natural biopolymer derived from chitin, has garnered significant attention owing to its potential chemical, biological, and physical properties, such as biocompatibility, bioactivity, and biosafety. This comprehensive review traces the historical development of CTS-based materials and delves into their specific applications across various fields. The study highlights the evolution of CTS from its initial discovery to its current state, emphasizing key milestones and technological advancements that have expanded its utility. Despite the extensive research, the synthesis and functionalization of CTS to achieve desired properties for targeted applications remain a challenge. This review addresses current problems such as the scalability of production, consistency in quality, and the environmental impact of extraction and modification processes. Additionally, it explores the novel applications of CTS-based materials in biomedicine, agriculture, environmental protection, and food industry, showcasing innovative solutions and future potentials. By providing a detailed analysis of the current state of CTS research and identifying gaps in knowledge, this review offers a valuable resource for researchers and industry professionals. The novelty of this work lies in its holistic approach, combining historical context with a forward-looking perspective on emerging trends and potential breakthroughs in the field of CTS-based functional materials. Therefore, this review will be helpful for readers by summarizing recent advances and discussing prospects in CTS-based functional materials.

Turning black soldier fly rearing by-products into valuable materials: Valorisation through chitin and chitin nanocrystals production.

The industry of insect-based proteins as feed and food products has been encountering a huge development since the last decade, and industrial-scale factories are now arising worldwide. Among all the species studied, Black Soldier Fly is one of the most promising and farmed. This rearing activity generates several by-products in the form of chitin-rich biomass that can be valorised to keep a virtuous production cycle embedded in the scope of the bioeconomy. Herein, we report the isolation of chitin and, for the first time, chitin nanocrystals (ChNCs) from all the BSF rearing by-products, i.e., moults (larval exuviae, puparium) and dead adults. Extraction yields, were dependent on the type of by-products and ranged from 5.8 % to 20.0 %, and the chemical structure of the extracts exhibited typical features of α-chitin, confirmed by FTIR, NMR, XRD and TGA analysis. Both STEM in SEM and AFM analysis confirmed the isolation of chitin nanocrystals presenting a rod-like morphology. The average nanocrystal height estimated by AFM ranged from 13 to 27 nm depending on the by-product sample. The following results highlighted the potential of BSF rearing by-products, promoting an approach to valorise those industrial waste and paving the way towards insect-based biorefinery.

Advances in extraction, utilization, and development of chitin/chitosan and its derivatives from shrimp shell waste.

Shrimp consumption is in great demand among the seafood used globally. However, this expansion has resulted in the substantial generation and disposal of shrimp shell waste. Through literature search, it has been observed that since 2020, global scholars have shown unprecedented interest in shrimp shell waste and its chitin/chitosan. However, these new insights lack corresponding and comprehensive summarization and analysis. Therefore, this article provides a detailed review of the extraction methods, applications, and the latest research developments on chitin/chitosan from shrimp shells, including micro-nano derivatives, from 2020 to the present. The results indicate that chemical extraction remains the primary technique for the extraction and preparation of chitin/chitosan from shrimp shells. With further refinement and development, adjusting parameters in the chemical extraction process or employing auxiliary techniques such as microwave and radiation enable the customization of target products with different characteristics (e.g., deacetylation degree, molecular weight, and degree of acetylation) according to specific needs. Additionally, in pursuit of environmentally friendly, efficient, and gentle extraction processes, recent research has shifted toward microbial fermentation and green solvent methods for chitin/chitosan extraction. Beyond the traditional antibacterial, film-forming, and encapsulation functionalities, research into the applications of chitosan in biomedical, food processing, new materials, water treatment, and adsorption fields is gradually deepening. Chitin/chitosan derivatives and their modified products have also been a focal point of research in recent years. However, with the rapid expansion, the future development of chitin/chitosan and its derivatives still faces challenges related to the unclear mechanism of action and the complexities associated with industrial scale-up.

Composite suture material based on polylactide accelerates the healing of surgical wounds in in vivo experiment.

In laboratory conditions, composite sutures based on polylactide (PLA) containing chitin nanofibrils modified with polyethylene glycol (CN-PEG) and poviargol (silver nanoparticles stabilized with poly(N-vinylpyrrolidone)) were obtained, studied, and used as a prototype. Surgical sutures threads with the addition of CN-PEG have stable mechanical properties both in air and in a buffer simulating the environment of a living organism. The yield strength of oriented threads decreased by an average of 15%, whereas for non-oriented threads the decrease was 3-4 times. The strength values in simple units of unfilled PLA, PLA containing 5 wt % CN-PEG, and PLA with 1 wt % Poviargol were on average 50% higher than the national standard 31620-2012. The results of in vivo experiments on albino rats (cross-linking skin and muscle tissue in the linea alba area) showed that composite sutures are best for suturing muscle tissue, whereas unfilled PLA sutures are more suitable for suturing skin. When suturing muscle tissue, suturing with composite sutures increased the number of collagen fibers of different diameters.

The beetle's structural protein CPCFC making elytra tough and rigid.

The insect cuticle, which serves as both a protective barrier and an efficient lever system for locomotion, is an extracellular matrix primarily composed of chitin and protein. The cuticle protein CPCFC characterized by a "CFC" motif containing 2 Cys split by the insertion of 5 residues is distributed across most insect species and specifically localized in the hard part of the cuticle. However, their physiological function is not fully understood. Here, we report 2 CPCFC proteins, TcCPCFC1 and TcCPCFC2, derived from the Coleopteran insect Tribolium castaneum. We revealed that TcCPCFC1 and TcCPCFC2 were predominantly expressed during the larval and adult stages of T. castaneum, respectively. The transcription downregulation of TcCPCFC1 significantly decreased the modulus and toughness of the elytral cuticle. We found that TcCPCFC proteins have high binding affinity to chitin. We cloned and produced recombinant TcCPCFC proteins and demonstrated that the addition of TcCPCFC proteins to chitin hydrogel greatly enhanced the hydrogel's modulus and toughness by forming denser chitin fibrous networks. Our findings reveal the functional role of CPCFC proteins in enhancing mechanical properties of insect cuticle, and we validate this process in vitro, and offer a protein candidate for fabrication of advanced chitin-based materials.

On the Valorisation of Chitin-Derived Furans by Milling.

Chitin-derived furans offer a sustainable alternative feedstock for nitrogen appended aromatic compounds. Herein, we address the challenge of using chitin-derived furans, 3-acetamido-5-acetylfuran (3A5AF) and 3-acetamido-5-furfural aldehyde (3A5F), to favour the formation of exo Diels-Alder adducts and 4­acetylaminophthalimides respectively, using a mechanochemical ball-milling technique. Mechanochemical activation is explored through the synthesis of 7-oxa-norbornene backbones with novel substitution pattern from 3A5AF in yields up to 77% and improved exo:endo selectivity compared to solution-phase reactions. The synthesis of 4­acetylaminophthalimides from 3A5F in yields up to 79% is also showcased from hydrazone derivatives.

Green extraction of chitin from hard spider crab shells.

A green protocol to extract chitin from crab shells using water soluble ionic liquids (ILs) is here reported. Compared to conventional multistep acid-base extraction methods, this one-pot procedure achieves pulping of recalcitrant crustacean waste shells by employing ammonium acetate, ammonium formate and hydroxylammonium acetate as water-soluble, low-cost and easy to prepare ILs. An extensive parametric analysis of the pulping process has been carried out with different ILs, different ratios, temperature and time. The optimized protocol provides a high-quality chitin comparable, if not better, to commercial chitin. The best results were obtained at 150 °C with ammonium formate prepared in-situ from aqueous ammonia and formic acid: chitin was isolated in a 17 wt% yield (based on dried crab shells as starting biowaste), a degree of acetylation (DA) > 94 %, a crystallinity index of 39-46 %, a molecular weight up to 6.6 × 105 g/mol and a polydispersity of ca 2.0.

Eumelanin-Enhanced Photothermal Disinfection of Contact Lenses Using a Sustainable Marine Nanoplatform Engineered with Electrospun Nanofibers.

Bacterial keratitis (BK) is a severe eye infection commonly associated with Staphylococcus aureus (S. aureus), posing a significant risk to vision, especially among contact lens wearers. This research introduces a novel smart nanoplatform (deMS@cNF), developed from demineralized mussel shells (deMS) and reinforced with chitin (CT) nanofibrils, specifically designed for portable photothermal disinfection of contact lenses. The nanoplatform leverages the photothermal properties of eumelanin in mussel shells (MS), which, when activated by a simple bike flashlight, rapidly heats to temperatures up to 95 °C, effectively destroying bacterial contamination. In vitro tests demonstrate that the nanoplatform is biocompatible and non-toxic, making it suitable for medical applications. This study highlights an innovative approach to converting marine biowaste into a safe, effective, and low-cost portable method for disinfecting contact lenses, showcasing the potential of the deMS@cNF platform for broader antimicrobial applications.

New Antibacterial and Antioxidant Chitin Derivatives: Ultrasonic Preparation and Biological Effects.

This work focuses on the first use of ultrasonic phenol-ene coupling as a polymer analogous transformation. The ultrasonic reaction was introduced into chitin chemistry, resulting in the fast and convenient preparation of new water-soluble cationic chitin derivatives. Since water-soluble derivatives of fully deacetylated chitin are poorly described in the literature, the synthesis of each new type of these derivatives is a significant event in polysaccharide chemistry. Polycations, or cationic polymers, are of particular interest as antibacterial agents. Consequently, the resulting polymers were tested for their antibacterial activity and toxicity. We found that the highly substituted polymer of medium molecular weight exhibited the most pronounced in vitro antibacterial effect. We prepared nanoparticles using the ionic gelation technique. The most effective in vitro antibacterial chitin-based systems were tested in vivo in rats. These tests demonstrated outstanding antibacterial effects combined with an absence of toxicity. Additionally, we found that the resulting polymers, unlike their nanoparticle counterparts, also exhibited strong antioxidant effects. In summary, we demonstrated the effectiveness of ultrasound in polymer chemistry and highlighted the importance of the sonochemical approach in the chemical modification of polysaccharides. This approach enables the synthesis of derivatives with improved physicochemical and biological properties.

Design, Synthesis, and Acaricidal Activity of 2,5-Diphenyl-1,3-oxazoline Compounds.

By using a scaffold hopping/ring equivalent and intermediate derivatization strategies, a series of compounds of 2,5-diphenyl-1,3-oxazoline with substituent changes at the 5-phenyl position were prepared, and their acaricidal activity was studied. However, the synthesized 2,5-diphenyl-1,3-oxazolines showed lower activity against mite eggs and larvae compared to the 2,4-diphenyl-1,3-oxazolines with the same substituents. We speculate that there is a significant difference in the spatial extension direction of the substituents between the two skeletons of compounds, resulting in differences in their ability to bind to the potential target chitin synthase 1. This work is helpful in inferring the internal structure of chitin synthase binding pockets.

Evolution of chitin-synthase in molluscs and their response to ocean acidification.

Chitin-synthase (CHS) is found in most eukaryotes and has a complex evolutionary history. Research into CHS has mainly been in the context of biomineralization of mollusc shells an area of high interest due to the consequences of ocean acidification. Exploration of CHS at the genomic level in molluscs, the evolution of isoforms, their tissue distribution, and response to environmental challenges are largely unknown. Exploiting the extensive molecular resources for mollusc species it is revealed that bivalves possess the largest number of CHS genes (12-22) reported to date in eukaryotes. The evolutionary tree constructed at the class level of molluscs indicates four CHS Type II isoforms (A-D) probably existed in the most recent common ancestor, and Type II-A (Type II-A-1/Type II-A-2) and Type II-C (Type II-C-1/Type II-C-2) underwent further differentiation. Non-specific loss of CHS isoforms occurred at the class level, and in some Type II (B-D groups) isoforms the myosin head domain, which is associated with shell formation, was not preserved and highly species-specific tissue expression of CHS isoforms occurred. These observations strongly support the idea of CHS functional diversification with shell biomineralization being one of several important functions. Analysis of transcriptome data uncovered the species-specific potential of CHS isoforms in shell formation and a species-specific response to ocean acidification (OA). The impact of OA was not CHS isoform-dependent although in Mytilus, Type I-B and Type II-D gene expression was down-regulated in both M. galloprovincialis and M. coruscus. In summary, during CHS evolution the gene family expanded in bivalves generating a large diversity of isoforms with different structures and with a ubiquitous tissue distribution suggesting that chitin is involved in many biological functions. These findings provide insight into CHS evolution in molluscs and lay the foundation for research into their function and response to environmental changes.

Enhanced degradation of insoluble chitin: Engineering high-efficiency chitinase fusion enzymes for sustainable applications.

N-acetyl-D-glucosamine and its dimer are degradation products of chitin waste with great potential in therapeutic and agricultural applications. However, the hydrolysis of insoluble chitin by chitinases remains a major bottleneck. This study investigated the biochemical properties and catalytic mechanisms of PoChi chitinase obtained from Penicillium oxalicum with a focus on enhancing its efficiency during the degradation of insoluble chitin. Recombinant plasmids were engineered to incorporate chitin-binding (ChBD) and/or fibronectin III (FnIII) domains. Notably, PoChi-FnIII-ChBD exhibited the highest substrate affinity (Km = 2.7 mg/mL) and a specific activity of 15.4 U/mg, which surpasses those of previously reported chitinases. These findings highlight the potential of engineered chitinases in advancing industrial biotechnology applications and offer a promising approach to more sustainable chitin waste management.

Intein Based Fusion Proteins: Great Tags for the Soluble Production and Convenient Purification of Recombinant Proteins.

Background: The main problem in the recombinant protein expression in E. coli strains, especially for high-yield production, is the accumulation in un-folded and inactive inclusion bodies. A suitable solution is the direction into the soluble cytoplasmic products by solubilizing tags. The use of inteins with self-cleaving ability, in addition to increase the chance of soluble protein expression, facilitates their purification process. Evidence Acquisition: In this review article, papers related to the use of intein tags for soluble expression or protein purification were collected regardless the time limit. Available databases including Pubmed, google scholar, ScienceDirect, Web of Science, Scopus, and Embase was searched. The best condition for soluble expression or purification was focused in all articles. Results: There are various intein tags commercially available in expression vectors that results in gaining our goal in facilitating the recombinant protein solubilization as well as its simple purification. It is enough to induce the self-cleavage property of the intein, which varies according to the type of intein used. In this way, the target protein is easily separated from the purification tag without the need to add protease enzymes such as enterokinase or treatment with various chemicals. The most common affinity tag in intein-based systems is Chitin Binding Domain attached to the chitin resin. Conclusions: In this review article, we introduced proteins or peptides which produced in fusion to intein tags and discussed about their expression condition and purification process in order to enhance the chance of soluble expression and intein cleavage in a single stage, respectively.

Valorization of fishery industry waste: Chitosan extraction and its application in the industry.

Waste from the fishing industry is disposed of in soils and oceans, causing environmental damage. However, it is also a source of valuable compounds such as chitin. Although chitin is the second most abundant polymer in nature, its use in industry is limited due to the lack of standardized and scalable extraction methods and its poor solubility. The deacetylation process increases its potential applications by enabling the recovery of chitosan, which is soluble in dilute acidic solutions. Chitosan is a polymer of great importance due to its biocompatible and bioactive properties, which include antimicrobial and antioxidant capabilities. Chitin extraction and its deacetylation to obtain chitosan are typically performed using chemical processes that involve large amounts of strongly acidic and alkaline solutions. To reduce the environmental impact of this process, extraction methods based on biotechnological tools, such as fermentation and chitin deacetylase, as well as emerging technologies, have been proposed. These extraction methods have demonstrated the potential to reduce or even avoid using strong solvents and shorten extraction time, thereby reducing costs. Nevertheless, it is important to address existing gaps in this area, such as the requirements for large-scale implementation and the determination of the stoichiometric ratios for each process. This review highlights the use of biotechnological tools and emerging technologies for chitin extraction and chitosan production. These approaches truly minimize environmental impact, reduce the use of strong solvents, and shorten extraction time. They are a reliable alternative to fishery waste valorization, lowering costs; however, addressing the critical gaps for their large-scale implementation remains challenging.