Chitin Synthesis and Degradation in Crustaceans: A Genomic View and Application

Chitin is among the most important components of the crustacean cuticular exoskeleton and intestinal peritrophic matrix. With the progress of genomics and sequencing technology, a large number of gene sequences related to chitin metabolism have been deposited in the GenBank database in recent years. Here, we summarized the genes and pathways associated with the biosynthesis and degradation of chitins in crustaceans based on genomic analyses. We found that chitin biosynthesis genes typically occur in single or two copies, whereas chitin degradation genes are all multiple copies. Moreover, the chitinase genes are significantly expanded in most crustacean genomes. The gene structure and expression pattern of these genes are similar to those of insects, albeit with some specific characteristics. Additionally, the potential applications of the chitin metabolism genes in molting regulation and immune defense, as well as industrial chitin degradation and production, are also summarized in this review.     

Chitin and Chitosan: Prospective Biomedical Applications in Drug Delivery,Cancer Treatment,and Wound Healing

Chitin and its derivative chitosan are highly abundant polymers in nature, appearing in both the shells and exoskeletons of various marine and non-marine species. Since they possess favorable properties, such as biocompatibility, biodegradability, non-toxicity, and non-immunogenicity, they have gained recent attention due to their enormous potential biomedical applications. The polycationic surface of chitosan enables it to form hydrogenic and ionic bonds with drug molecules, which is one of its most useful properties. Because chitosan is biocompatible, it can therefore be used in drug delivery systems. The development of chitosan-based nanoparticles has also contributed to the significance of chitin as a drug delivery system that can deliver drugs topically. Furthermore, chitin can be used in cancer treatment as a vehicle for delivering cancer drugs to a specific site and has an antiproliferative effect by reducing the viability of cells. Finally, chitosan can be used as a wound dressing in order to promote the faster regeneration of skin epithelial cells and collagen production by fibroblasts. As discussed in this review, chitin and chitosan have diverse applications in the medical field. Recognizing the biomedical applications of these two polymers is essential for future research in tissue engineering and nanobiotechnology.     

Chitin and Chitosan as Multipurpose Natural Polymers for Groundwater Arsenic Removal and As2O3 Delivery in Tumor Therapy

Chitin and chitosan are natural polysaccharide polymers. These polymers have been used in several agricultural, food protection and nutraceutical applications. Moreover, chitin and chitosan have been also used in biomedical and biotechnological applications as drug delivery systems or in pharmaceutical formulations. So far, there are only few studies dealing with arsenic (As) removal from groundwater using chitin or chitosan and no evidence of the use of these natural polymers for arsenic trioxide (As₂O₃) delivery in tumor therapy. Here we suggest that chitin and/or chitosan might have the right properties to be employed as efficient polymers for such applications. Besides, nanotechnology offers suitable tools for the fabrication of novel nanostructured materials of natural origin. Since different nanostructured materials have already been employed successfully in various multidisciplinary fields, we expect that the integration of nanotechnology and natural polymer chemistry will further lead to innovative applications for environment and medicine.     

N-acetylglucosamine: production and applications

N-Acetylglucosamine (GlcNAc) is a monosaccharide that usually polymerizes linearly through (1,4)-β-linkages. GlcNAc is the monomeric unit of the polymer chitin, the second most abundant carbohydrate after cellulose. In addition to serving as a component of this homogeneous polysaccharide, GlcNAc is also a basic component of hyaluronic acid and keratin sulfate on the cell surface. In this review, we discuss the industrial production of GlcNAc, using chitin as a substrate, by chemical, enzymatic and biotransformation methods. Also, newly developed methods to obtain GlcNAc using glucose as a substrate in genetically modified microorganisms are introduced. Moreover, GlcNAc has generated interest not only as an underutilized resource but also as a new functional material with high potential in various fields. Here we also take a closer look at the current applications of GlcNAc, and several new and cutting edge approaches in this fascinating area are thoroughly discussed.     

Chitin Deacetylases: Properties and Applications

Chitin deacetylases, occurring in marine bacteria, several fungi and a few insects, catalyze the deacetylation of chitin, a structural biopolymer found in countless forms of marine life, fungal cell and spore walls as well as insect cuticle and peritrophic matrices. The deacetylases recognize a sequence of four GlcNAc units in the substrate, one of which undergoes deacetylation: the resulting chitosan has a more regular deacetylation pattern than a chitosan treated with hot NaOH. Nevertheless plain chitin is a poor substrate, but glycolated, reprecipitated or depolymerized chitins are good ones. The marine Vibrio sp. colonize the chitin particles and decompose the chitin thanks to the concerted action of chitinases and deacetylases, otherwise they could not tolerate chitosan, a recognized antibacterial biopolymer. In fact, chitosan is used to prevent infections in fishes and crustaceans. Considering that chitin deacetylases play very important roles in the biological attack and defense systems, they may find applications for the biological control of fungal plant pathogens or insect pests in agriculture and for the biocontrol of opportunistic fungal human pathogens.     

Modification of Chitin with Kraft Lignin and Development of New Biosorbents for Removal of Cadmium(II) and Nickel(II) Ions

Novel, functional materials based on chitin of marine origin and lignin were prepared. The synthesized materials were subjected to physicochemical, dispersive-morphological and electrokinetic analysis. The results confirm the effectiveness of the proposed method of synthesis of functional chitin/lignin materials. Mechanism of chitin modification by lignin is based on formation of hydrogen bonds between chitin and lignin. Additionally, the chitin/lignin materials were studied from the perspective of waste water treatment. The synthetic method presented in this work shows an attractive and facile route for producing low-cost chitin/lignin biosorbents with high efficiency of nickel and cadmium adsorption (88.0% and 98.4%, respectively). The discovery of this facile method of synthesis of functional chitin/lignin materials will also have a significant impact on the problematic issue of the utilization of chitinous waste from the seafood industry, as well as lignin by-products from the pulp and paper industry.     

Emerging Biomedical Applications of Nano-Chitins and Nano-Chitosans Obtained via Advanced Eco-Friendly Technologies from Marine Resources

The present review article is intended to direct attention to the technological advances made in the 2010–2014 quinquennium for the isolation and manufacture of nanofibrillar chitin and chitosan. Otherwise called nanocrystals or whiskers, n-chitin and n-chitosan are obtained either by mechanical chitin disassembly and fibrillation optionally assisted by sonication, or by e-spinning of solutions of polysaccharides often accompanied by poly(ethylene oxide) or poly(caprolactone). The biomedical areas where n-chitin may find applications include hemostasis and wound healing, regeneration of tissues such as joints and bones, cell culture, antimicrobial agents, and dermal protection. The biomedical applications of n-chitosan include epithelial tissue regeneration, bone and dental tissue regeneration, as well as protection against bacteria, fungi and viruses. It has been found that the nano size enhances the performances of chitins and chitosans in all cases considered, with no exceptions. Biotechnological approaches will boost the applications of the said safe, eco-friendly and benign nanomaterials not only in these fields, but also for biosensors and in targeted drug delivery areas.     

Chitosan: An Update on Potential Biomedical and Pharmaceutical Applications

Chitosan is a natural polycationic linear polysaccharide derived from chitin. The low solubility of chitosan in neutral and alkaline solution limits its application. Nevertheless, chemical modification into composites or hydrogels brings to it new functional properties for different applications. Chitosans are recognized as versatile biomaterials because of their non-toxicity, low allergenicity, biocompatibility and biodegradability. This review presents the recent research, trends and prospects in chitosan. Some special pharmaceutical and biomedical applications are also highlighted.     

Design of Chitosan and Its Water Soluble Derivatives-Based Drug Carriers with Polyelectrolyte Complexes

Chitosan, the cationic polysaccharide derived from the natural polysaccharide chitin, has been studied as a biomaterial for more than two decades. As a polycationic polymer with favorable properties, it has been widely used to form polyelectrolyte complexes with polyanions for various applications in drug delivery fields. In recent years, a growing number of studies have been focused on the preparation of polyelectrolyte complexes based on chitosan and its water soluble derivatives. They have been considered well-suited as biomaterials for a number of vital drug carriers with targeted/controlled release profiles, e.g., films, capsules, microcapsules. In this work, an overview highlights not only the favorable properties of chitosan and its water soluble derivatives but also the good performance of the polyelectrolyte complexes produced based on chitosan. Their various types of applications as drug carriers are reviewed in detail.     

Chitosan Modification and Pharmaceutical/Biomedical Applications

Chitosan has received much attention as a functional biopolymer for diverse applications, especially in pharmaceutics and medicine. Our recent efforts focused on the chemical and biological modification of chitosan in order to increase its solubility in aqueous solutions and absorbability in the in vivo system, thus for a better use of chitosan. This review summarizes chitosan modification and its pharmaceutical/biomedical applications based on our achievements as well as the domestic and overseas developments: (1) enzymatic preparation of low molecular weight chitosans/chitooligosaccharides with their hypocholesterolemic and immuno-modulating effects; (2) the effects of chitin, chitosan and their derivatives on blood hemostasis; and (3) synthesis of a non-toxic ion ligand—D-Glucosaminic acid from Oxidation of D-Glucosamine for cancer and diabetes therapy.     

Antimicrobial properties of chitosan nanoparticles: Mode of action and factors affecting activity

The present investigation describes the synthesis and characterization of novel biodegradable nanoparticles based on chitosan for biomedical applications. The presence of primary amine groups in repeating units of chitosan grants it several properties like antibacterial activity, antitumor activity and so on. Chitosan forms nanoparticles spontaneously on the addition of polyanion tripolyphosphate which has greater antimicrobial activity than parent chitosan. In the present study, chitosan nanoparticles (ChNP) were prepared by the ionic gelation method. The physiochemical characteristics of nanoparticles were analyzed using XRD, SEM, FTIR. The antibacterial activity of chitosan nanoparticles against medical pathogens Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa was evaluated by calculation of minimum inhibitory concentration (MIC) and compared with chitosan and chitin activity. The mode of action and factors affecting antibacterial activity were also analyzed. ChNP compounds exhibited superior antimicrobial activity against all microorganisms in comparison with chitosan and chitin. The antibiofilm activity was studied using crystal violet assay and growth on congo red agar. The study is thus a good demonstration of the applicability of chitosan nanoparticles as an effective antimicrobial agent with antibiofilm activity as well.     

Biodegradation and Prospect of Polysaccharide from Crustaceans

Marine crustacean waste has not been fully utilized and is a rich source of chitin. Enzymatic degradation has attracted the wide attention of researchers due to its unique biocatalytic ability to protect the environment. Chitosan (CTS) and its derivative chitosan oligosaccharides (COSs) with various biological activities can be obtained by the enzymatic degradation of chitin. Many studies have shown that chitosan and its derivatives, chitosan oligosaccharides (COSs), have beneficial properties, including lipid-lowering, anti-inflammatory and antitumor activities, and have important application value in the medical treatment field, the food industry and agriculture. In this review, we describe the classification, biochemical characteristics and catalytic mechanisms of the major degrading enzymes: chitinases, chitin deacetylases (CDAs) and chitosanases. We also introduced the technology for enzymatic design and modification and proposed the current problems and development trends of enzymatic degradation of chitin polysaccharides. The discussion on the characteristics and catalytic mechanism of chitosan-degrading enzymes will help to develop new types of hydrolases by various biotechnology methods and promote their application in chitosan.     

Chitin Degradation Machinery and Secondary Metabolite Profiles in the Marine Bacterium Pseudoalteromonas rubra S4059

Genome mining of pigmented Pseudoalteromonas has revealed a large potential for the production of bioactive compounds and hydrolytic enzymes. The purpose of the present study was to explore this bioactivity potential in a potent antibiotic and enzyme producer, Pseudoalteromonas rubra strain S4059. Proteomic analyses (data are available via ProteomeXchange with identifier PXD023249) indicated that a highly efficient chitin degradation machinery was present in the red-pigmented P. rubra S4059 when grown on chitin. Four GH18 chitinases and two GH20 hexosaminidases were significantly upregulated under these conditions. GH19 chitinases, which are not common in bacteria, are consistently found in pigmented Pseudoalteromonas, and in S4059, GH19 was only detected when the bacterium was grown on chitin. To explore the possible role of GH19 in pigmented Pseudoalteromonas, we developed a protocol for genetic manipulation of S4059 and deleted the GH19 chitinase, and compared phenotypes of the mutant and wild type. However, none of the chitin degrading ability, secondary metabolite profile, or biofilm-forming capacity was affected by GH19 deletion. In conclusion, we developed a genetic manipulation protocol that can be used to unravel the bioactive potential of pigmented pseudoalteromonads. An efficient chitinolytic enzyme cocktail was identified in S4059, suggesting that this strain could be a candidate with industrial potential.     

“绿肥绿药美土嘉禾”农田生态新技术

生物熏蒸就是利用芸苔属植物细胞破碎后硫苷与黑芥子酶接触反应生成的挥发性异硫氰酸盐／酯类,熏蒸土壤、杀灭／抑制土生病虫害。该文摘编2008年在堪培拉召开的第三届国际生物熏蒸学术会议的22个国家和地区的77篇论文以及21期生物熏蒸最新报道,综述在高发病虫害农田,间、套、偷种、复种或轮作高硫苷油／芥菜品种,适时适量碎汁＋入土＋捂青,确是一项绿肥绿药美土嘉禾的农田生态新技术。中国农田之广、耕制之多,在果菜园、棉薯田、城郊集约种植基地,试验、示范推广绿肥绿药美土嘉禾农田生态新技术必将大利归农、惠及全民的。     

Proteases Production and Chitin Preparation from the Liquid Fermentation of Chitinous Fishery By-Products by Paenibacillus elgii

Chitinous fishery by-products have great application in the production of various bioactive compounds. In this study, Paenibacillus elgii TKU051, a protease-producing bacterial strain, was isolated using a medium containing 1% squid pens powder (SPP) as the sole carbon/nitrogen (C/N) source. P. elgii TKU051 was found to produce at least four proteases with molecular weights of 100 kDa, 57 kDa, 43 kDa, and 34 kDa (determined by the gelatin zymography method). A P. elgii TkU051 crude enzyme cocktail was optimally active at pH 6–7 and 60 °C. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and α-glucosidase inhibitory activity of the hydrolysates obtained from the hydrolysis of shrimp shell powder, shrimp head powder, shrimp meat powder, fish head powder and soya bean powder catalyzed by the P. elgii TkU051 crude enzyme cocktail were also evaluated. P. elgii TKU051 exhibited a high deproteinization capacity (over 94%) on different kinds of shrimp waste (shrimp heads and shells; fresh and cooked shrimp waste; shrimp waste dried by oven and lyophilizer), and the Fourier-transform infrared spectroscopy profile of the chitin obtained from the deproteinization process displayed the characteristic of chitin. Finally, the obtained chitin exhibited an effect comparable to commercial chitin in terms of adsorption against Congo Red (90.48% and 90.91%, respectively). Thus, P. elgii TKU051 showed potential in the reclamation of chitinous fishery by-products for proteases production and chitin extraction.     

Bioproduction of Chitooligosaccharides: Present and Perspectives

Chitin and chitosan oligosaccharides (COS) have been traditionally obtained by chemical digestion with strong acids. In light of the difficulties associated with these traditional production processes, environmentally compatible and reproducible production alternatives are desirable. Unlike chemical digestion, biodegradation of chitin and chitosan by enzymes or microorganisms does not require the use of toxic chemicals or excessive amounts of wastewater. Enzyme preparations with chitinase, chitosanase, and lysozymeare primarily used to hydrolyze chitin and chitosan. Commercial preparations of cellulase, protease, lipase, and pepsin provide another opportunity for oligosaccharide production. In addition to their hydrolytic activities, the transglycosylation activity of chitinolytic enzymes might be exploited for the synthesis of desired chitin oligomers and their derivatives. Chitin deacetylase is also potentially useful for the preparation of oligosaccharides. Recently, direct production of oligosaccharides from chitin and crab shells by a combination of mechanochemical grinding and enzymatic hydrolysis has been reported. Together with these, other emerging technologies such as direct degradation of chitin from crustacean shells and microbial cell walls, enzymatic synthesis of COS from small building blocks, and protein engineering technology for chitin-related enzymes have been discussed as the most significant challenge for industrial application.     

Identification and Characterization of Three Chitinases with Potential in Direct Conversion of Crystalline Chitin into N,N′-diacetylchitobiose

Chitooligosaccharides (COSs) have been widely used in agriculture, medicine, cosmetics, and foods, which are commonly prepared from chitin with chitinases. So far, while most COSs are prepared from colloidal chitin, chitinases used in preparing COSs directly from natural crystalline chitin are less reported. Here, we characterize three chitinases, which were identified from the marine bacterium Pseudoalteromonas flavipulchra DSM 14401^T, with an ability to degrade crystalline chitin into (GlcNAc)₂ (N,N’-diacetylchitobiose). Strain DSM 14401 can degrade the crystalline α-chitin in the medium to provide nutrients for growth. Genome and secretome analyses indicate that this strain secretes six chitinolytic enzymes, among which chitinases Chia4287, Chib0431, and Chib0434 have higher abundance than the others, suggesting their importance in crystalline α-chitin degradation. These three chitinases were heterologously expressed, purified, and characterized. They are all active on crystalline α-chitin, with temperature optima of 45–50 °C and pH optima of 7.0–7.5. They are all stable at 40 °C and in the pH range of 5.0–11.0. Moreover, they all have excellent salt tolerance, retaining more than 92% activity after incubation in 5 M NaCl for 10 h at 4 °C. When acting on crystalline α-chitin, the main products of the three chitinases are all (GlcNAc)₂, which suggests that chitinases Chia4287, Chib0431, and Chib0434 likely have potential in direct conversion of crystalline chitin into (GlcNAc)₂.     

Chitosan, the marine functional food, is a potent adsorbent of humic acid

Chitosan is prepared by the deacetylation of chitin, the second-most abundant biopolymer in nature, and has applicability in the removal of dyes, heavy metals and radioactive waste for pollution control. In weight-reduction remedies, chitosan is used to form hydrogels with lipids and to depress the intestinal absorption of lipids. In this study, an experimental method was implemented to simulate the effect of chitosan on the adsorption of humic acid in the gastrointestinal tract. The adsorption capacity of chitosan was measured by its adsorption isotherm and analyzed using the Langmuir equation. The results showed that 3.3 grams of humic acid was absorbed by 1 gram of chitosan. The adsorption capacity of chitosan was much greater than that of chitin, diethylaminoethyl-cellulose or activated charcoal. Cellulose and carboxymethyl-cellulose, a cellulose derivative with a negative charge, could not adsorb humic acid in the gastrointestinal tract. This result suggests that chitosan entraps humic acid because of its positive charge.     

A Broad-Specificity Chitinase from Penicillium oxalicum k10 Exhibits Antifungal Activity and Biodegradation Properties of Chitin

Penicillium oxalicum k10 isolated from soil revealed the hydrolyzing ability of shrimp chitin and antifungal activity against Sclerotinia sclerotiorum. The k10 chitinase was produced from a powder chitin-containing medium and purified by ammonium sulfate precipitation and column chromatography. The purified chitinase showed maximal activity toward colloidal chitin at pH 5 and 40 °C. The enzymatic activity was enhanced by potassium and zinc, and it was inhibited by silver, iron, and copper. The chitinase could convert colloidal chitin to N-acetylglucosamine (GlcNAc), (GlcNAc)2, and (GlcNAc)3, showing that this enzyme had endocleavage and exocleavage activities. In addition, the chitinase prevented the mycelial growth of the phytopathogenic fungi S. sclerotiorum and Mucor circinelloides. These results indicate that k10 is a potential candidate for producing chitinase that could be useful for generating chitooligosaccharides from chitinous waste and functions as a fungicide.