Structure of insect chitin isolated from beetle larva cuticle and silkworm (Bombyx mori) pupa exuvia

Chitin samples in a alpha-form structure were isolated from beetle larva cuticle and silkworm (Bombyx mori) pupa exuvia by treatment with 1 N HCl and 1 N NaOH. Chitosan was prepared by treating them in 40% NaOH containing NaBH(4). Chitin and chitosan were analyzed by X-ray, [13C]CP/MAS NMR, [13C]FT-NMR, and scanning electron microscopy (SEM) methods. Insect chitin degraded more readily than shrimp chitin when treated with 6 N HCl and the enzyme-chitinase. After treatment with 2 N HCl at 100 degrees C, the insect chitin crystallinity increased. N-deacetylation of insect chitin was easier than that of crustaceous chitin, and about 94% of the N-acetyl groups were removed in one treatment with 40% NaOH for 4 h at 110 degrees C. After treatment with 2 N HCl, 55% of the N-acetyl groups of silkworm chitin were removed under the same conditions. Beetle chitin showed a higher affinity for chitinase than shrimp chitin.     

Study on the production of chitin and chitosan from shrimp shell by using Bacillus subtilis fermentation

Fermentation of shrimp shell in jaggery broth using Bacillus subtilis for the production of chitin and chitosan was investigated. It was found that B. subtilis produced sufficient quantities of acid to remove the minerals from the shell and to prevent spoilage organisms. The protease enzyme in Bacillus species was responsible for the deprotenisation of the shell. The pH, proteolytic activity, extent of demineralization and deprotenisation were studied during fermentation. About 84% of the protein and 72% of the minerals were removed from the shrimp shell after fermentation. Mild acid and alkali treatments were given to produce characteristic chitin and their concentrations were standardized. Chitin was converted to chitosan by N-deacetylation and the properties of chitin and chitosan were studied. FTIR spectral analysis of chitin and chitosan prepared by the process was carried out and compared with spectra of commercially available samples.     

Applications of magnetic resonance spectroscopy to chitin from insect cuticles

Chitin is the second most abundant polysaccharide in nature after cellulose. At the present time, the main commercial sources of chitin are the crab and shrimp shells which are major waste products from the seafood industry. However, current chitin resources have some inherent problems including seasonal availability, limited supplies, and environmental pollution. As an alternative, insect cuticle is proposed as an unconventional but viable source of chitin. This review focuses on the recent sources of insect chitin and the application of various magnetic resonance spectroscopic techniques to native insect cuticles, particularly cicada sloughs and chitin extracted from insect sloughs. In addition, the physicochemical properties, isolation process, and degree of N-acetylation (DA) is reviewed and discussed.     

Chitin and chitosan from Basidiomycetes

Chitinous material was isolated from the mycelium of seven species of Basidiomycetes to evaluate the possibility of using fungal biomass as a source of chitin and chitosan. Such material was characterised for its purity, degree of acetylation and crystallinity. Chitin yields ranged between 8.5 and 19.6% dry weight and the chitosan yield was approximately 1%. The characteristics of the fungal chitins were similar to those of commercial chitin. Chitosans, with a low degree of acetylation, comparable with that of commercial chitosan, were obtained by the chemical deacetylation of fungal chitins.     

Deacetylation of Chitin under Homogeneous Conditions

Chitin isolated enzymatically from Antarctic krill shells was dissolved in aqueous NaOH by freezing and thawing to create homogeneous conditions. Deacetylation was performed at room temperature or under heating. The degree of deacetylation, molecular weight, and dynamic viscosity of solutions were estimated in chitosan samples. Deacetylation of chitin under homogeneous conditions was optimized. Chitosans with molecular weights of 180–220 and 250–300 kDa were obtained from the chitins of Antarctic krill and northern shrimp, respectively.     

Lime treatment of shrimp head waste for the generation of highly digestible animal feed

In addition to approximately 20% ash, shrimp processing by-products contain 64% protein and chitin, both of which can be used to generate several valuable products. Chitin and chitosan production is currently based on several crustacean wastes, and at the present time the protein fraction is not being used. This paper describes the thermo-chemical treatment of shrimp processing wastes with lime to generate a protein-rich material with a well-balanced amino acid content that can be used as a monogastric animal feed supplement. The residual solid, rich in calcium carbonate and chitin, can still be used to generate chitin and chitosan through well-established processes.     

Removal of copper, chromium, and arsenic from CCA-treated wood onto chitin and chitosan

Chitin and chitosan are naturally abundant biopolymers which are of interest to research concerning the sorption of metal ions since the amine and hydroxyl groups on their chemical structures act as chelation sites for metal ions. This study evaluates the removal of copper, chromium, and arsenic elements from chromated copper arsenate (CCA)-treated wood via biosorption by chitin and chitosan. Exposing CCA-treated sawdust to various amounts of chitin and chitosan for 1, 5, and 10 days enhanced removal of CCA components compared to remediation by deionized water only. Remediation with a solution containing 2.5 g chitin for 10 days removed 74% copper, 62% chromium, and 63% arsenic from treated sawdust. Remediation of treated sawdust samples using the same amount of chitosan as chitin resulted in 57% copper, 43% chromium, and 30% arsenic removal. The results suggest that chitin and chitosan have a potential to remove copper element from CCA-treated wood. Thus, these more abundant natural amino polysaccharides could be important in the remediation of waste wood treated with the newest formulations of organometallic copper compounds and other water-borne wood preservatives containing copper.     

Extraction and Characterization of α-Chitin and Chitosan from Six Different Aquatic Invertebrates

Chitin and chitosan were extracted from six different aquatic invertebrate species. Species dry weights varied between 5 % and 20 % chitin, and the chitosan productivity of these chitins varied between 66 % and 74 %. Chitin and chitosan structures were characterized by FTIR, TGA, XRD, and SEM. FTIR results showed that the chitins obtained from the organisms were observed in α form. Chitin thermal stabilities were in the order Ranatra linearis?>?Anax imperator?>?Hydrophilus piceus?>?Notonecta glauca?>?Agabus bipustulatus?>?Asellus aquaticus, and chitosan thermal stabilities in the order N. glauca?>?A. bipustulatus?>?A. imperator?>?R. linearis?>?H. piceus?>?A. aquaticus. The crystalline index values of chitins varied between 76.4 % and 90.6 %. Their surface morphology was examined by SEM, revealing nanofibre structures. These six aquatic invertebrate species with characterized chitin and chitosan structures may be used as alternative chitin and chitosan sources for various technological purposes.     

Highly Fibrous and Porous Raw Material Shaped Chitin Isolated from <Emphasis Type="Italic">Oniscus</Emphasis> sp. (Crustacea)

Chitin was isolated from a crustacean by keeping the original body shape for the first time. The isolation method followed in this study was simple and time and energy saving unlike the labor-intensive classical methods. Chitin samples preserving the original shape were isolated from Isopoda (Oniscus sp.) successfully in a total of 20 min including filtration time. FT-IR, XRD and TGA analysis and chitinase digestion test demonstrated that the chitin was pure, low crystalline (Crystalline index: 51 %) and had low thermal stability (maximum degradation temperature: 328.8 °C). SEM analysis revealed the highly fibrous structure of chitin. The chitin content of the whole body was found significantly higher (29.6?±?4.2 %) than the earlier reports. Interaction of chitin isolates with Bovine Serum Albumin protein were studied at different pH. It was concluded that this three dimensional raw material shaped chitin can be effectively used in any adsorption studies due to 1) highly fibrous and porous nature, 2) low crystallinity and 3) low thermal stability. And also this biodegradable and biocompatible biopolymer can be suggested as a carrier matrix in further studies thanks to its three dimensional shape.     

Effect of Explosion on the Crystalline Polymorphism of Chitin and Chitosan

For identification of how explosion increases the reactivity of chitin and chitosan, changes in the crystalline polymorphism of these polysaccharides were studied by X-ray diffraction measurements. The α-chitin form of chitin did not change after being exploded, but an X-ray diagram of chitosan showed a hydrated crystal of low crystallinity before the explosion, and increased crystallinity of the hydrated form plus a small amount of an anhydrous crystal after the explosion. The improvement of accessibility to both polysaccharides caused by the explosion seemed not to arise from changes in their crystalline polymorphism or crystallinity     

Extraction and characterization of chitin and chitosan from marine sources in Arabian Gulf

Chitin in the α and the β forms has been extracted from different marine crustacean from the Arabian Gulf. The contents of the various exoskeletons have been analyzed and the percent of the inorganic salt (including the various elements present), protein and the chitin was determined. Deacetylation of the different chitin produced was conducted by the conventional thermal heating and by microwave heating methods. Microwave heating has reduced enormously the time of heating from 6–10 h to 10–15 min, to yield the same degree of deacetylation and higher molecular weight chitosan. This technique can save massive amount of energy when implemented on a semi-industrial or industrial scale. The chitin and the obtained chitosan were characterized by elemental analysis, XRD, NMR, FTIR and thermogravimetric measurements. XRD analysis showed that chitosan has lower crystallinity than its corresponding chitin; meanwhile its thermal stability is also lower than chitin.     

Evaluation of chitosans and Pichia guillermondii as growth inhibitors of Penicillium digitatum

Chitosans were obtained by room-temperature-homogeneous-deacetylation (RTHD) and freeze-pump-out-thaw-heterogeneous-deacetylation (FPT) from chitins purified from fermentations. Commercial chitosan was deacetylated by three-FPT-cycles. Chitosans and Pichia guillermondii were evaluated on the growth of Penicillium digitatum. Medium molecular weight (M(W)) chitosans displayed higher inhibitory activity against the yeast than low M(W) biopolymers. Chitosans with low degree of acetylation (DA) were inhibitory for yeast and mould. Therefore, a low M(W) and high DA chitosan was selected for use against moulds combined with yeasts. Biopolymer and yeasts presented an additive effect, since chitosans were effective to delay spore germination, whereas yeast decreased apical fungal growth.     

Mining marine shellfish wastes for bioactive molecules: chitin and chitosan--Part A: extraction methods

Legal restrictions, high costs and environmental problems regarding the disposal of marine processing wastes have led to amplified interest in biotechnology research concerning the identification and extraction of additional high grade, low-volume by-products produced from shellfish waste treatments. Shellfish waste consisting of crustacean exoskeletons is currently the main source of biomass for chitin production. Chitin is a polysaccharide composed of N-acetyl-D-glucosamine units and the multidimensional utilization of chitin derivatives including chitosan, a deacetylated derivative of chitin, is due to a number of characteristics including: their polyelectrolyte and cationic nature, the presence of reactive groups, high adsorption capacities, bacteriostatic and fungistatic influences, making them very versatile biomolecules. Part A of this review aims to consolidate useful information concerning the methods used to extract and characterize chitin, chitosan and glucosamine obtained through industrial, microbial and enzymatic hydrolysis of shellfish waste.     

Extraction and characterization of chitin and chitosan from local sources

Chitin has been extracted from six different local sources in Egypt. The obtained chitin was converted into the more useful soluble chitosan by steeping into solutions of NaOH of various concentrations and for extended periods of time, then the alkali chitin was heated in an autoclave which dramatically reduced the time of deacetylation. Chitin from squid pens did not require steeping in sodium hydroxide solution and showed much higher reactivity towards deacetylation in the autoclave that even after 15 min of heating a degree of deacetylation of 90% was achieved. The obtained chitin and chitosan were characterized by spectral analysis, X-ray diffraction and thermo gravimetric analysis.     

Peripheral enzymatic deacetylation of chitin and reprecipitated chitin particles

The enzymatic deacetylation of various chitin preparations was investigated using the fungal chitin deacetylase (CDA) isolated from Rhizopus oryzae growth medium. Specific extracellular enzyme activity after solid state fermentation was 10 times higher than that after submerged fermentation. Natural crystalline chitin is a very poor substrate for the enzyme, but showed a five-time better deacetylation after dissolution and reprecipitation. Chitin particles, enzymatically deacetylated for only 1% exhibited a strongly increased binding capacity towards ovalbumin, while maintaining the rigidity and insolubility of chitin in a moderate acidic environment. Because of the unique combination of properties, these CDA treated chitin materials were named "chit-in-osan". Chitinosan was shown to be an attractive matrix for column chromatography because no hydrogel formation was observed, that impaired the flow of eluent. Under the same conditions, partially deacetylated chitosan swelled and blocked the flow in the column.     

Physicochemical behavior of homogeneous series of acetylated chitosans in aqueous solution: role of various structural parameters

Physicochemical properties of four different homogeneous series of chitosans with degrees of acetylation (DA) and weight-average degrees of polymerization (DP(w)) ranging from 0 to 70% and 650 to 2600, respectively, were characterized in an ammonium acetate buffer (pH 4.5). Then, the intrinsic viscosity ([eta](0)), the root-mean-square z-average of the gyration radius (R(G,z)), and the second virial coefficient (A(2)) were studied by viscometry and static light scattering. The conformation of chitosan, according to DA and DP(w), was highlighted through the variations of alpha and nu parameters, deduced from the scale laws [eta](0) = K(w)and R(G,z) = K', respectively, and the total persistence length (L(p,tot)). In relation with the different behaviors of chitosan in solution, the conformation varied according to two distinct domains versus DA with a transition range in between. Then, (i) for DA < 25%, chitosan exhibited a flexible conformation; (ii) a transition domain for 25 < DA < 50%, where the chitosan conformation became slightly stiffer and, (iii) for DA > 50%, on increasing DP(w) and DA, the participation of the excluded volume effect became preponderant and counterbalanced the depletion of the chains by steric effects and long-distance interactions. It was also highlighted that below and beyond a critical DP(w,c) (ranging from 1 300 to 1 800 for DAs from 70 to 0%, respectively) the flexibility of chitosan chains markedly increased then decreased (for DA > 50%) or became more or less constant (DA < 50%). All the conformations of chitosan with regards to DA and DP(w) were described in terms of short-distance interactions and excluded volume effect.     

Functional characteristics of shrimp chitosan and its membranes as affected by the degree of deacetylation

The functional properties of three shrimp chitosan preparations with different degrees of deacetylation (75%, 87% and 96% DD) but with a constant molecular weight (about 810 kDa) were investigated. Chitosan with 75% DD had a 1.5 times higher water absorption, probably due to its 20% lower level of crystallinity. Membranes cast from this chitosan also exhibited 1.5 times more water absorption and 2 times higher permeability. However, chitosan with 87% and 96% DD had 1.5-2 times higher absorption of fat and the orange II dye. This is attributed to the higher content of positively charged amine groups in the polymer. Cast into membrane, chitosan of higher degree of deacetylation showed a higher tensile strength and a higher elongation at break, probably due to the higher level of crystallinity.     

Studies on applications of chitin and its derivatives

Chitin, a homopolymer of N-acetylglucosamine, is obtained from a variety of sources. They form the structural component of fungal cell wall and plants. They are commercially obtained from shrimp and crab shell waste from the fishing industry. Recent advances in understanding the structure and properties of chitin and its derivatives has opened a lot of new avenues for its applications. Improvements in the properties of chitin for a particular application can be easily brought about by chemical modifications. The applicability of chitin in many areas and its easy manipulation has resulted in a considerable amount of research being done on the possible applications of chitinase.     

The extracellular constitutive production of chitin deacetylase in Metarhizium anisopliae: possible edge to entomopathogenic fungi in the biological control of insect pests

The possible contribution of extracellular constitutively produced chitin deacetylase by Metarhizium anisopliae in the process of insect pathogenesis has been evaluated. Chitin deacetylase converts chitin, a beta-1,4-linked N-acetylglucosamine polymer, into its deacetylated form chitosan, a glucosamine polymer. When grown in a yeast extract-peptone medium, M. anisopliae constitutively produced the enzymes protease, lipase, and two chitin-metabolizing enzymes, viz. chitin deacetylase (CDA) and chitosanase. Chitinase activity was induced in chitin-containing medium. Staining of 7.5% native polyacrylamide gels at pH 8.9 revealed CDA activity in three bands. SDS-PAGE showed that the apparent molecular masses of the three isoforms were 70, 37, and 26 kDa, respectively. Solubilized melanin (10microg) inhibited chitinase activity, whereas CDA was unaffected. Following germination of M. anisopliae conidia on isolated Helicoverpa armigera, cuticle revealed the presence of chitosan by staining with 3-methyl-2-benzothiazoline hydrazone. Blue patches of chitosan were observed on cuticle, indicating conversion of chitin to chitosan. Hydrolysis of chitin with constitutively produced enzymes of M. anisopliae suggested that CDA along with chitosanase contributed significantly to chitin hydrolysis. Thus, chitin deacetylase was important in initiating pathogenesis of M. anisopliae softening the insect cuticle to aid mycelial penetration. Evaluation of CDA and chitinase activities in other isolates of Metarhizium showed that those strains had low chitinase activity but high CDA activity. Chemical assays of M. anisopliae cell wall composition revealed the presence of chitosan. CDA may have a dual role in modifying the insect cuticular chitin for easy penetration as well as for altering its own cell walls for defense from insect chitinase.     

Solid-state characterization of chitosans derived from lobster chitin

Two samples of chitosan (CH1 and CH2) of different molecular weights and degrees of deacetylation were prepared from lobster chitin under two different processes. Solid-state properties of CH1 and CH2 were characterized and compared with four commercial chitosans prepared from crab and fresh shrimp shells. Infrared spectroscopy (IR), solid-state CP-MAS ¹³C NMR, powder X-ray diffraction and differential scanning calorimetric techniques were used to characterize the molecular structure and solid-state properties of the materials. Changes in the crystallinity and polymorphic forms of CH1 and CH2 were attributable to the different process conditions used. The differences in crystallinity were confirmed by powder X-ray diffraction data. The methods of preparation of CH1 and CH2 did not significantly influence the bulk, tap and true densities of the bulk material, but they affected the flow properties of CH1 and CH2. In conclusion, the physicochemical properties of the present chitosans prepared from lobster chitin (CH1 and CH2) are comparable with those of commercial chitosan materials of crab or shrimp shell origin.