Optimization of Dothideomycetes sp. css035 chitosanase productivity and activity using response surface methodology (original) (raw)
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Applied Sciences, 2021
Chitooligosaccharides (COS) are homo- or hetero-oligomers of D-glucosamine (GlcN) and N-acetyl-D-glucosamine (GlcNAc) that can be obtained by chitosan or chitin hydrolysis. Their enzymatic production is preferred over other methodologies (physical, chemical, etc.) due to the mild conditions required, the fewer amounts of waste and its efficiency to control product composition. By properly selecting the enzyme (chitinase, chitosanase or nonspecific enzymes) and the substrate properties (degree of deacetylation, molecular weight, etc.), it is possible to direct the synthesis towards any of the three COS types: fully acetylated (faCOS), partially acetylated (paCOS) and fully deacetylated (fdCOS). In this article, we review the main strategies to steer the COS production towards a specific group. The chemical characterization of COS by advanced techniques, e.g., high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and MALDI-TOF mass spectrometry,...
Production and Biological Activities of Chitooligosaccharides (COS)-An Overview
Journal of Chitin and Chitosan Science, 2015
The food, pharmaceutical and cosmeceutical industries have been actively involved in the exploitation of chitooligosac-charides (COS) in various formulations, as they possess unmatched biological activities. In this review we report the marine resources of chitosan and the methods in the production of COS with various methods including chemical and enzymatic methods. In addition, we have made an attempt to report the latest research on the biological activities of COS such as antioxidation, antiinflammation, antimicrobial, anticancer and anti-HIV effects. The reports suggest that COS have more efficacy in controlling pathological responses in humans. The biological effects of the COS could prove as beneficial inputs for the biotechnological industries to use them as leads in various formulations.
Production of Chitooligosaccharides and Their Potential Applications in Medicine
Marine Drugs, 2010
Chitooligosaccharides (CHOS) are homo-or heterooligomers of N-acetylglucosamine and D-glucosamine. CHOS can be produced using chitin or chitosan as a starting material, using enzymatic conversions, chemical methods or combinations thereof. Production of well-defined CHOS-mixtures, or even pure CHOS, is of great interest since these oligosaccharides are thought to have several interesting bioactivities. Understanding the mechanisms underlying these bioactivities is of major importance. However, so far in-depth knowledge on the mode-of-action of CHOS is scarce, one major reason being that most published studies are done with badly characterized heterogeneous mixtures of CHOS. Production of CHOS that are well-defined in terms of length, degree of N-acetylation, and sequence is not straightforward. Here we provide an overview of techniques that may be used to produce and characterize reasonably well-defined CHOS fractions. We also present possible medical applications of CHOS, including tumor growth inhibition and inhibition of T H 2-induced inflammation in asthma, as well as use as a bone-strengthener in osteoporosis, a vector for gene delivery, an antibacterial agent, an antifungal agent, an anti-malaria agent, or a hemostatic agent in wound-dressings. By using well-defined CHOS-mixtures it will become possible to obtain a better understanding of the mechanisms underlying these bioactivities.
Chitooligosaccharides (COS) is a derived oligosaccharides mixture produced from chitosan. COS possess various bioactivity of pharmacological importance. Enzymatic conversion of chitosan yields COS of defined degree of polymerization, rather than acid hydrolysis. In this study, chitosanase producing microorganisms, mainly bacteria was isolated from marine soil. About 25 isolates were screened for chitosanase production based on quantitative determination of reducing sugar using DNSA method. High enzyme activity exhibiting isolate was identified using 16S rDNA gene amplification, based on BLAST analysis, the organism was found to be Bacillus firmus. The enzyme production was maximum during 72 h incubation in static mode. The maximum enzyme activity was obtained at pH 9, temperature 50˚C. Thin Layer chromatography revealed the maximal enzymatic production of COS was observed during 3-7 h. Hence, this enzyme could be exploited for the production of COS.
International Journal of Pharmacy and Pharmaceutical Sciences, 2024
Objective: The present study was carried out to develop an enzymatic hydrolysate with unique biological properties targeting diabetic foot ulcers. Methods: Chitosanase-producing organisms were isolated and used to create chitooligosaccharide hydrolysate. Various techniques, such as FTIR, NMR, and X-ray diffraction, were used. Antimicrobial activity was tested using disc diffusion and well diffusion methods. Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were determined through the Chitooligosaccharide-Broth Dilution Method. Results: The study identified marine mud samples and isolated S9, S15, and SF12 as significant sources of chitosanase production. The partially purified chitosanolytic enzymes produced by these isolates were hydrolyzed in a 1% chitosan solution at 180 °C, revealing more prominent antimicrobial activity. The Chitooligosaccharide Hydrolysate (COS) preparation was fixed at 45 °C, pH 5.5, for 180 min. The chitosanase enzyme was soluble in four solvents and insoluble in ethanol, acetone, and diethyl ether. All COS hydrolysates prepared showed antimicrobial activity against foot ulcer pathogens, Pseudomonas sp., and Candida albicans. S9 COS showed higher activity than SF12 hydrolysates against foot ulcer pathogens. The COS hydrolysate showed significantly stronger antimicrobial activities than chitosan and chitosanase. Conclusion: The present study concludes that COS hydrolysate and its biological functions are applicable for diabetic foot ulcer treatment. Further investigation into the efficacy of COS against diverse infectious pathogens is needed.
Chitooligosaccharides (COS) are -1,4-linked homo-oligosaccharides of N-acetylglucosamine (GlcNAc) or glucosamine (GlcN), and also include hetero-oligosaccharides composed of GlcNAc and GlcN. These sugars are of practical importance because of their various biological activities, such as anti-microbial, anti-inflammatory, anti-oxidant and anti-tumor activities, as well as triggering the innate immunity in plants. The reported data on bioactivities of COS used to contain some uncertainties or contradictions, because the experiments were conducted with poorly characterized COS mixtures. Recently, COS have been satisfactorily characterized with respect to their structures, especially the degree of polymerization (DP) and degree of N-acetylation (DA); thus, the structure-bioactivity relationship of COS has become more unambiguous. To date, various green-chemical strategies involving enzymatic synthesis of COS with designed sequences and desired biological activities have been developed....
Biochemical Engineering Journal, 2005
Chitosan was depolymerized either by HCl hydrolysis or enzymatic degradation with a commercial preparation Pectinex Ultra Spl. The chitooligosaccharides released by both methods were selectively precipitated in methanol solutions and characterized using MALDI-TOF mass spectrometry. Differences between the two methods were detected and concerned the degrees of polymerization of the fragments produced and their acetylation. The enzymatic method yielded shorter fragments with a higher proportion of fully deacetylated chitooligomers. Conversely, acid hydrolysis of the starting chitosan resulted in fragments with degrees of polymerization up to sixteen and more monoacetylated residues than with the enzymatic procedure.
Journal of Agricultural and Food Chemistry, 2013
Chitooligosaccharides possessing remarkable biological properties can be obtained by enzymatic hydrolysis of chitin. In this work, the chitosanase activity of soluble and immobilized glycosyltransferase (Branchzyme ®) toward chitosan, as well as its biochemical characterization are described for the first time. This enzyme was found to be homotetrameric with a mol. wt. of 256 kDa, an isoelectric point of 5.3, and an optimal temperature range of between 50-60°C. It was covalently immobilized to glutaraldehyde-agarose with protein and activity immobilization yields of 67% and 17% respectively. Immobilization improved enzyme stability, increasing its half life fivefold, and allowed enzyme reuse for at least 25 consecutive cycles. The chitosanase activity of Branchzyme ® on chitosan was similar for the soluble and immobilized forms. The reaction mixture was constituted by chitooligosaccharides with degrees of polymerization (DP) of between 2 to 20, with a higher concentration having degrees of polymerization of 3 to 8.
Chitosanases produced by microbes and plants are getting attention to explore vastly available marine waste. Chitooligosaccharides and glucosamine can be produced using chitosanase enzyme and have applications in food, pharma and other industries. A potential microbial chitosanase source was found after isolation and screening of chitosan degrading microbes from garden soil. An isolate, designated as C6 produced chitosanase enzyme upon induction by chitosan substrates. Production of 6 U/ml of chitosanase enzyme was achieved from this isolate on chitosan minimal salt broth medium at 32°C after 3 days of growth. The enzyme was able to hydrolyse both chitosan and cellulosic substrates. Enzymatic production of D-glucosamine and chitooligosaccharides were studied with various chitosan substrates using crude enzyme. The yield of glucosamine was found to be 40% after 2 h of reaction at 40°C, and chitosan oligomers were produced having two to six polymerizations at 60°C reaction temperature. The hydrolysates showed 50% antioxidant activity as compared to ascorbic acid.
Preparation of chitooligosaccharides from fungal waste mycelium by recombinant chitinase
Carbohydrate research, 2016
This study aimed to develop an enzymatic method for conversion of chitin from fungal waste mycelia to chitooligosaccharides. The recombinant chitinase LlChi18A from Lactococcus lactis was over-expressed by Escherichia coli BL21 (DE3) and purified by affinity chromatography. The enzymatic properties of the purified enzyme were studied by chitin oligosaccharides. Waste mycelium was pre-treated by alkaline. The optimal conditions for hydrolysis of fungal chitin by recombinant chitinase were determined by Schales method. HPLC/ESI-MS was used to determine the content of N-acetylglucosamine and chitooligosaccharides after hydrolysis. The level of reducing sugar released from pretreated mycelium by chitinase increased with the reaction time during 6 days. The main product in the hydrolysates was N,N'-diacetylchitobiose. After hydrolysis by chitinase for 5 d, the yield of N,N'-diacetylchitobiose from waste mycelium was around 10% with estimated purity of around 70%. Combination of c...