Synthesis and characterization of phosphorylcholine-substituted chitosans soluble in physiological pH conditions

A polymer analogous synthesis involving the reductive amination of phosphorylcholine (PC)-glyceraldehyde with primary amines of deacetylated chitosan (M(w) approximately 57000 g mol(-1)) was used to prepare phosphorylcholine-substituted chitosans (PC-CH) with a degree of substitution (DS) ranging from approximately 11 to approximately 53 mol % PC-substituted glucosamine residues. The PC-CH derivatives were characterized by (1)H NMR spectroscopy, FTIR spectroscopy, and multiangle laser light scattering gel permeation chromatography (MALLS-GPC). The pK(a) of the PC-substituted amine groups (pK(a) approximately 7.20) was determined by (1)H NMR titration. The PC-CH samples (1.0 g L(-1)) were shown to be nontoxic using an MTT assay performed with human KB cells. Aqueous solutions of PC-CH samples (4.0 g L(-1)) of DS >or= 22 mol % PC-substituted glucosamine residues remained clear, independently of pH (4.0 < pH < 11.0). The remarkable water solubility and nontoxicity displayed by the new PC-CH samples open up new opportunities in the design of chitosan-based biomaterials and nanoparticles.     

N-(carboxymethylidene)chitosans and N-(carboxymethyl)chitosans: Novel chelating polyampholytes obtained from chitosan glyoxylate

Glyoxylic acid, added to aqueous suspensions of chitosan, causes immediate dissolution of chitosan and gel formation within 3–4 h if the pH is 4.5–5.5. Solutions at lower pH values gel after 2 min of warming at 60–80°. Chitosan glyoxylate solutions brought to alkaline pH with sodium hydroxide do not precipitate chitosan. Evidence is given that a Schiff base, namely N-(carboxymethylidene)chitosan, is formed. N-(Carboxymethylidene)chitosans are reduced by sodium cyanoborohydride at room temperature to give N-(carboxymethyl)chitosans, obtained as white, free-flowing powders, soluble in water at all pH values. A series of N-(carboxymethyl)chitosans having various degrees of acetylation and N-carboxymethylation was obtained, and characterized by viscometry, elemental analysis, and i.r. spectrometry. For the fully substituted N-(carboxymethyl)chitosans, the pK′ is 2.3, the pK″ is 6.6, and the isoelectric point is 4.1. The addition of N-(carboxymethyl)chitosan to solutions (0.2–0.5mm) of transition-metal ions produces immediate insolubilization of N-(carboxymethyl)chitosan-metal ion chelates.     

Characterization and antimicrobial activity of water-soluble N-(4-carboxybutyroyl) chitosans against some plant pathogenic bacteria and fungi

Water-soluble N-(4-carboxybutyroyl) chitosan derivatives with different degrees of substitution (DS) were synthesized to enhance the antimicrobial activity of chitosan molecule against plant pathogens. Chitosan in a solution of 2% aqueous acetic acid-methanol (1:1, v/v) was reacted with 0.1, 0.3, 0.6 and 1 mol of glutaric anhydride to give N-(4-carboxybutyroyl) chitosans at DS of 0.10, 0.25, 0.48 and 0.53, respectively. The chemical structures and DS were characterized by ¹H and ¹³C NMR spectroscopy, which showed that the acylate reaction took place at the N-position of chitosan. The synthesized derivatives were more soluble than the native chitosan in water and in dilute aqueous acetic acid and sodium hydroxide solutions. The antimicrobial activity was in vitro investigated against the most economic plant pathogenic bacteria of Agrobacterium tumefaciens and Erwinia carotovora and fungi of Botrytis cinerea, Pythium debaryanum and Rhizoctonia solani. The antimicrobial activity of N-(4-carboxybutyroyl) chitosans was strengthened than the un-modified chitosan with the increase of the DS. A compound of DS 0.53 was the most active one with minimum inhibitory concentration (MIC) of 725 and 800 mg/L against E. carotovora and A. tumefaciens, respectively and also in mycelial growth inhibiation against B. cinerea (EC₅₀ = 899 mg/L), P. debaryanum (EC₅₀ = 467 mg/L) and R. solani (EC₅₀ = 1413 mg/L).     

Synthesis,characterization and antifungal activity of quaternary derivatives of chitosan on Aspergillus flavus

Two series of new chitosan derivatives were synthesized by reaction of deacetylated chitosan (CH) with propyl (CH-Propyl) and pentyl (CH-Pentyl) trimethylammonium bromides to obtain derivatives with increasing degrees of substitution (DS). The derivatives were characterized by ¹H NMR and potentiometric titration techniques and their antifungal activities on the mycelial growth of Aspergillus flavus were investigated in vitro. The antifungal activities increase with DS and the more substituted derivatives of both series, CH-Propyl and CH-Pentyl, exhibited antifungal activities respectively three and six times higher than those obtained with commercial and deacetylated chitosan. The minimum inhibitory concentrations (MIC) were evaluated at 24, 48 and 72 h by varying the polymer concentration from 0.5 to 16 g/L and the results showed that the quaternary derivatives inhibited the fungus growth at polymer concentrations four times lower than that obtained with deacetylated chitosan (CH). The chitosans modified with pentyltrimethylammonium bromide exhibited higher activity and results are discussed taking into account the degree of substitution (DS).     

Synthesis of cis-(1-->3)-glycosides of allyl 2-acetamido-4,6-O-benzylidene-2-deoxy-alpha-D-glucopyranoside

Syntheses of allyl 2,3,4-tri-O-benzyl-alpha-D-gluco- and D-galactopyranosyluronate-(1-->3)-2-acetamido-4,6-O-benzylidene-2-deoxy-alpha-D-glucopyranoside via oxidation of the hydroxymethyl group of allyl 2,3,4-tri-O-benzyl-alpha-D-gluco- and D-galactopyranosyl-(1-->3)-2-acetamido-4,6-O-benzylidene-2-deoxy-alpha-D-glucopyranoside under Jones conditions are described. Structures of the title compounds were confirmed by (1)H and (13)C NMR spectroscopy.     

Determination of the degree of N-acetylation and the distribution of N-acetyl groups in partially N-deacetylated chitins (chitosans) by high-field n.m.r. spectroscopy.

The composition and sequence of 2-acetamido-2-deoxy-beta-D-glucose (GlcNAc) and 2-amino-2-deoxy-beta-D-glucose (GlcN) residues in partially N-deacetylated chitosans, prepared under homogeneous and heterogeneous conditions, have been determined by 1H-n.m.r. spectroscopy. It was necessary to depolymerise the chitosan slightly by treatment with nitrous acid before spectroscopy. A sequence-dependent deshielding of H-1 of the GlcNAc residues made it possible to determine the proportions of the four possible diads. Chitosan prepared by N-deacetylation under homogeneous conditions gave values for the diad frequencies that were roughly consistent with a random distribution of the N-acetyl groups. Samples prepared under heterogeneous conditions have a frequency of the GlcNAc-GlcNAc diad slightly higher than for a random (Bernoullian) distribution. The chitosans, prepared under both homogeneous and heterogeneous conditions, with a degree of acetylation of 50% were soluble at neutral pH.     

Physicochemical characterization of chitin and chitosan from crab shells

Crab chitosan was prepared by alkaline N-deacetylation of crab chitin for 60, 90 and 120 min and the yields were 30.0-32.2% with that of chitosan C120 being the highest. The degree of N-deacetylation of chitosans (83.3–93.3%) increased but the average molecular weight (483–526 kDa) decreased with the prolonged reaction time. Crab chitosans showed lower lightness and WI values than purified chitin, chitosans CC and CS but higher than crude chitin. With the prolonged reaction time, the nitrogen (8.9–9.5%), carbon (42.2–45.2%) and hydrogen contents (7.9–8.6%) in chitosans prepared consistently increased whereas N/C ratios remained the same (0.21). Crab chitosans prepared showed a melting endothermic peak at 152.3–159.2 °C. Three chitosans showed similar microfibrillar crystalline structure and two crystalline reflections at 2θ = 8.8–9.0° and 18.9–19.1°. Overall, the characteristics of three crab chitosans were unique and differed from those of chitosan CC and CS as evidenced by the element analysis, differential scanning calorimetry, scanning electron microscopy and X-ray diffraction patterns.     

Copolymers of ethylene imine and N-(2-hydroxyethyl)-ethylene imine as tools to study effects of polymer structure on physicochemical and biological properties of DNA complexes

A series of five poly[(ethylene imine)-co-N-(2-hydroxyethyl-ethylene imine)] copolymers with similar molecular weights and different degrees of branching was established to study structure-function relationship with regard to physicochemical and biological properties as gene delivery systems. Copolymers were synthesized by acid-catalyzed ring-opening copolymerization of aziridine and N-(2-hydroxyethyl)-aziridine in aqueous solution and characterized by GPC-MALLS, (1)H- and (13)C NMR, IR, potentiometric titration, and ion exchange chromatography. Complexation of DNA was determined by agarose gel electrophoresis, and complex sizes were quantitated by PCS. Cytotoxicity of the copolymers in fibroblasts was assessed by MTT-assay, LDH-assay, and hemolysis. The transfection efficiency was determined using the reporter plasmid pGL3 in 3T3 mouse fibroblasts. The copolymers obtained by solution polymerization had relatively low molecular weights of about 2000 Da, and the degree of branching increased with increasing ethylene imine ratio. The pK(a) as well as the buffer capacity increased proportional to the number of primary and secondary amines. Higher branched polymers showed stronger complexation and condensation of DNA, formed smaller polymer/DNA complexes, and induced the expression of plasmids to a higher extent than less branched polymers. In vitro cytotoxic effects and the hemolysis of erythrocytes decreased with decreased branching. Our results indicate that the basicity and degree of protonation of the polymers depending on their amount of primary and secondary amines seem to be important factors both for their transfection efficiency and for their cytotoxicity in gene transfer.     

Chemical modification of chitosan. Part 15: synthesis of novel chitosan derivatives by substitution of hydrophilic amine using N-carboxyethylchitosan ethyl ester as an intermediate

The Michael type reaction of chitosan with ethyl acrylate has been investigated. Although this reaction was quite slow in the case of chitosan, the reiteration of the reaction was an effective means for increasing the degree of substitution (DS) of ethyl ester. The N-carboxyethylchitosan ethyl ester as an intermediate was successfully substituted with various hydrophilic amines, although the simultaneous hydrolysis of the ester to carboxylic acid also occurred. Water-soluble chitosan derivatives were obtained by substitution with hydroxyalkylamines and diamines.     

Evidence of two-step deprotonation of D-mannitol in aqueous solution

Deprotonation of D-mannitol was studied in aqueous basic solutions by means of potentiometry and (13)C NMR spectroscopy. Two-step dissociation in the pH range from 12 to 13.8 was shown, and successive dissociation constants K(a1) and K(a2) were determined. In a solution with ionic strength I = 1.0 M (NaOH + NaNO(3)) pK(a1) = 13.1 +/- 0.1 and pK(a2) = 13.8 +/- 0.2. With increasing ionic strength from 0.75 to 3.0 M, both pK(a1) and pK(a2) values decrease. Deprotonation-induced chemical shifts in pH-variable (13)C NMR spectra show that the OH-groups next to internal carbon atoms C-3 and C-4 dissociate to a greater extent compared to OH-groups next to external carbon atoms C-1 and C-6.     

High yield production of monomer-free chitosan oligosaccharides by pepsin catalyzed hydrolysis of a high deacetylation degree chitosan

The high molecular weight of chitosan, which results in a poor solubility at neutral pH values and high viscosity aqueous solutions, limits its potential uses in the fields of food, health and agriculture. However, most of these limitations are overcome by chitosan oligosaccharides obtained by enzymatic hydrolysis of the polymer. Several commercial enzymes with different original specificities were assayed for their ability to hydrolyze a 93% deacetylation degree chitosan and compared with a chitosanase. According to the patterns of viscosity decrease and reducing end formation, three enzymes--cellulase, pepsin and lipase A--were found to be particularly suitable for hydrolyzing chitosan at a level comparable to that achieved by chitosanase. Unlike the appreciable levels of both 2-amino-2-deoxy-D-glucose and 2-acetamido-2-deoxy-D-glucose monomers released from chitosan by the other enzymes after a 20h-hydrolysis (4.6-9.1% of the total product weight), no monomer could be detected following pepsin cleavage. As a result, pepsin produced a higher yield of chitosan oligosaccharides than the other enzymes: 52% versus as much as 46%, respectively. Low molecular weight chitosans accounted for the remaining 48% of hydrolysis products. The calculated average polymerization degree of the products released by pepsin was around 16 units after 20h of hydrolysis. This product pattern and yield are proposed to be related to the bond cleavage specificity of pepsin and the high deacetylation degree of chitosan used as substrate. The optimal reaction conditions for hydrolysis of chitosan by pepsin were 40 degrees C and pH 4.5, and an enzyme/substrate ratio of 1:100 (w/w) for reactions longer than 1h.     

Theoretical prediction of relative and absolute pKa values of aminopyridines

This work presents a study aimed at the theoretical prediction of pK(a) values of aminopyridines, as a factor responsible for the activity of these compounds as blockers of the voltage-dependent K(+) channels. To cover a large range of pK(a) values, a total of seven substituted pyridines is considered as a calibration set: pyridine, 2-aminopyridine, 3-aminopyridine, 4-aminopyridine, 2-chloropyridine, 3-chloropyridine, and 4-methylpirydine. Using ab initio G1, G2 and G3 extrapolation methods, and the CPCM variant of the Polarizable Continuum Model for solvation, we calculate gas phase and solvation free energies. pK(a) values are obtained from these data using a thermodynamic cycle for describing protonation in aqueous and gas phases. The results show that the relatively inexpensive G1 level of theory is the most accurate at predicting pK(a) values in aminopyridines. The highest standard deviation with respect to the experimental data is 0.69 pK(a) units for absolute values calculations. The difference increases slightly to 0.74 pK(a) units when the pK(a) is computed relative to the pyridine molecule. Considering only compounds at least as basic as pyridine (the values of interest for bioactive aminopyridines) the error falls to 0.10 and 0.12 pK(a) units for the absolute and relative computations, respectively. The technique can be used to predict the effect of electronegative substituents in the pK(a) of 4-AP, the most active aminopyridine considered in this work. Thus, 2-chloro and 3-chloro-4-aminopyridine are taken into account. The results show a decrease of the pK(a), suggesting that these compounds are less active than 4-AP at blocking the K(+) channel.     

N-(2-Carboxyethyl)chitosans: regioselective synthesis,characterisation and protolytic equilibria

N-(2-Carboxyethyl)chitosans were obtained by reaction of low molecular weight chitosan with a low degree of acetylation and 3-halopropionic acids under mild alkaline media (pH 8-9, NaHCO3) at 60 degrees C. The chemical structure of the derivatives obtained was determined by 1H and 13C NMR spectroscopies. It was found that alkylation of chitosan by 3-halopropionic acids proceeds exclusively at the amino groups. The products obtained are described in terms of their degrees of carboxyethylation and ratio of mono-, di-substitution and free amine content. The protonation constants of amino and carboxylate groups of a series of N-(2-carboxyethyl)chitosans were determined by pH-titration at ionic strength 0.1 M KNO3 and 25 degrees C.     

The secondary cell wall polymer of Geobacillus tepidamans GS5-97T: structure of different glycoforms

Nuclear magnetic resonance spectroscopic studies of the strain-specific secondary cell wall polymer (SCWP) of the Gram-positive, moderately thermophilic organism Geobacillus tepidamans GS5-97T reveal two glycoforms consisting of identical tetrasaccharide repeating units with different chemical modifications of the amide moieties. On the basis of sugar analyses along with 1D and 2D 1H, 13C, 15N, and 31P NMR spectroscopy at natural isotope abundance, the basic backbone structure of the SCWP was established to be [beta-D-Manp-2,3-diNAcANH2-(1-->6)-alpha-D-Glcp-(1-->4)-beta-D-Manp-2,3-diNAcANH2-(1-->3)-alpha-D-GlcpNAc-(1-->]6-(1-->O)-PO2-(O-->6)-MurNAc-, with modifications of the amide groups. In one glycoform, all beta-D-Manp-2,3-diNAcANH2 (2,3-diacetamido-2,3-dideoxy-beta-D-mannopyranuronamide, ManpANH2) residues are substituted with two acetyl groups (glycoform I) at the amide group at C-6; in the other glycoform (glycoform II), only one proton of this amide group is substituted by an acetyl group. The ratio between both the glycoforms approximates 1:1.     

Chemoenzymatic synthesis of the 3-sulfated Lewisa pentasaccharide

The sulfated pentasaccharide benzyl O-(3-O-sulfo-beta-D-galactopyranosyl)-(1-->3)-O-[(alpha-L-fucopyranosyl)-(1-->4)]-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-(1-->3)-O-(beta-D-galactopyranosyl)-(1-->4)-O-beta-D-glucopyranoside sodium salt was synthesized using a chemo-enzymatic approach. Lacto-N-tetraose, obtained from two disaccharides [4-methoxybenzyl O-(2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1-->3)-4,6-O-benzylidene-2-deoxy-2-phtalimido-beta-D-glucopyranoside and benzyl 2,6-di-O-acetyl-beta-D-galactopyranosyl-(1-->4)-2,3,6-tri-O-acetyl-beta-D-glucopyranoside], was regioselectively sulfated at the 3 OH position of the terminal galactose using the stannylene procedure. The fucosylation of the sulfated tetrasaccharide was performed using soluble or immobilized fucosyltransferase FucT-III to give the title compound.     

Preparation of N-vanillyl chitosan and 4-hydroxybenzyl chitosan and their physico-mechanical, optical, barrier, and antimicrobial properties

Chitosan derivatives such as N-vanillyl chitosan and 4-hydroxybenzyl chitosan were prepared by reacting chitosan with 4-hydroxy-3-methoxybenzaldehyde (vanillin) and 4-hydroxybenzaldehyde. Amino groups on chitosan reacts with these aldehydes to form a Schiff base intermediate, which is later on converted into N-alkyl chitosans by reduction with sodium cyanoborohydride. The chemical reaction was monitored by ¹H NMR spectroscopy and the absence of aldehydic proton at 9.83 ppm in NMR spectra was observed for both the modified chitosan derivatives confirming the reaction. Modified chitosan films were later prepared by solution casting method and their physico-mechanical, barrier, optical and thermal properties were studied. The results clearly indicated significant change in tensile strength, water vapour transmission rate, and haze properties of modified chitosans. Modified chitosan films were also studied for their antimicrobial activity against Aspergillus flavus. The results showed a marked reduction of aflatoxins produced by the fungus in the presence of the N-vanillyl chitosan and 4-hydroxybenzyl chitosan film discs to 98.9% and non-detectable levels, respectively.     

Triterpenoid saponins with N-acetyl sugar from the bark of Albizia procera

Three (1,2,4) and one known (3) triterpenoid saponins were isolated from the bark of Albizia procera. The saponins were characterized as 3-O-[beta-D-xylopyranosyl-(1-->2)-alpha-L-arabinopyranosyl-(1-->6)-2-acetamido-2-deoxy-beta-D-glucopyranosyl] echinocystic acid (1), 3-O-[alpha-L-arabinopyranosyl-(1-->2)-beta-D-fucopyranosyl-(1-->6)-2-acetamido-2-deoxy-beta-D-glucopyranosyl] echinocystic acid (2) and 3-O-[beta-D-xylopyranosyl-(1-->2)-alpha-L-arabinopyranosyl-(1-->6)-2-acetamido-2-deoxy-beta-D-glucopyranosyl] acacic acid lactone (4). Their structures were elucidated by 1D and 2D NMR experiments, FABMS as well as chemical means. Saponins 1 and 3 exhibited cytotoxicity against HEPG2 cell line with IC50 9.13 microg/ml and 10 microg/ml, respectively.     

Effect of chitosans on in vitro rumen digestion and fermentation of maize silage

The gas in vitro technique was used to study the effects of six types of chitosans, each having different molecular weights and acetylation degrees, on rumen microbial fermentation. In a first trial, a separate concentration of 750 mg/l of culture fluid for each of the six chitosans (CHI1, CHI2, CHI3, CHI4, CHI5, and CHI6) was incubated for 24 h in diluted ruminal fluid with maize silage as the substrate. The ionophore antibiotic monensin (MON) was used as a positive control, and a negative control with no chitosan (CTR) was also included. Each treatment was tested in triplicate for three different periods. At the end of the trial, samples were collected to determine volatile fatty acid (VFA) and ammonia N concentrations, and pH and gas production values were recorded. Methane concentration was estimated stoichiometrically. In vitro true organic matter digestibility (IVOMD) and partitioning factor (PF, mg OM truly degraded/ml gas produced) were also calculated. In a second trial, a separate concentration of 750 mg/l of each of the six chitosans was incubated for 144 h in diluted ruminal fluid with maize silage as the substrate, to study the effects of these compounds on fermentation kinetics.All six chitosans decreased the IVOMD and PF values. Chitosan inclusion did not affect the fermentation of the substrate's soluble fraction, but did reduce the fermentation kinetics of the insoluble but fermentable fraction. However, only CHI5 and CHI6 decreased total VFA concentration. CHI3 and CHI6 decreased the molar proportion of acetate and increased the molar proportion of propionate, thus increasing the propionate-to-acetate ratio. Chitosan inclusion did not affect molar proportions of butyrate. With the exception of CHI2, the molar proportion of branch-chained VFA was lowered by all of the chitosan treatments. Most of the treatments also decreased methane production, also with the exception of CHI2.In conclusion, chitosan extracts may enable the manipulation of rumen microbial fermentation, but further research is required to elucidate the effect of chitosans on ruminal fermentation parameters in commercial diets as well as the adaptability of rumen microflora to these additives.     

Structure of a highly substituted beta-xylan of the gum exudate of the palm Livistona chinensis (Chinese fan)

Structural features of the acidic, highly substituted glycanoxylan (LCP; 87% yield) from the gum exudate of the palm, Livistona chinensis, family Arecaceae, were determined. It had [alpha]D -30 degrees, Mw 1.9x10(5) and a polydispersity ratio Mw/Mn of approximately 1.0. Acid hydrolysis gave rise to Rha, Fuc, Ara, Xyl, and Gal, in a 1:6:46:44:3 molar ratio, and 12% of uronic acid was present. LCP had a highly branched structure with side-chains containing nonreducing end-units (% values are approximate) of Araf (15%), Fucp (4%), Xylp (7%), GlcpA, and 4-Me-GlcpA, and internal 2-O- (5%) and 3-O-substituted Araf (8%), and 2-O-substituted Xylp (14%) units. The (1-->4)-linked beta-Xylp main-chain units of LCP were substituted at O-3 (4%), O-2 (17%), and O-2,3 (16%). Partial acid hydrolysis gave 4-Me-alpha-GlcpA-(1-->2)-[beta-Xylp-(1-->4)](0-2)-Xyl, identified by showing that the uronic acids were single-unit side-chain substituents on O-2. Milder hydrolysis conditions removed from O-3 other side-chains containing Fucp and Araf nonreducing end-units and internal Arap, and 2-O- and 3-O-substituted Araf units. Carboxyl-reduced LCP contained 4-O-methylglucose and glucose in a 3.2:1 molar ratio, arising from GlcpA and 4-OMe-GlcpA nonreducing end-units, respectively. The gum contained small amounts of free alpha-Fucp-(1-->2)-Ara, which corresponds to structures in the polysaccharide. Free myo- and D- or L-chiro-inositol were present in a 9:1 ratio.     

Relevance of molecular weight of chitosan and its derivatives and their antioxidant activities in vitro

The antioxidant potency of different molecular weight (DMW) chitosan and sulfated chitosan derivatives was investigated employing various established in vitro systems, such as superoxide (O(2)(.-))/hydroxyl ((-.)OH) radicals scavenging, reducing power, iron ion chelating. As expected, we obtained several satisfying results, as follows: firstly, low molecular weight chitosan had stronger scavenging effect on O(2)(.-) and (-.)OH than high molecular weight chitosan. For example the O(2)(.-) scavenging activity of low molecular weight chitosan (9 kDa) and high molecular weight chitosan (760 kDa) were 85.86% and 35.50% at 1.6 mg/mL, respectively. Secondly, comparing with DMW chitosan, DMW sulfated chitosans had the stronger inhibition effect on O(2)(.-). At 0.05 mg/mL, the scavenging activity on O(2)(.-) reached 86.26% for low molecular weight chitosan sulfate (9 kDa), but that of low molecular weight chitosan (9 kDa) was 85.86% at 1.6 mg/mL. As concerning chitosan and sulfated chitosan of the same molecular weight, scavenging activities of sulfated chitosan on superoxide and hydroxyl radicals were more pronounced than that of chitosan. Thirdly, low molecular weight chitosan sulfate had more effective scavenging activity on O(2)(.-) and (-.)OH than that of high molecular weight chitosan sulfate. Fourthly, DMW chitosans and sulfated chitosans were efficient in the reducing power, especially LCTS. Their orders were found to be LCTS>CTS4>HCTS>CTS3>CTS2>CTS1>CTS. Fifthly, CTS4 showed more considerable ferrous ion-chelating potency than others. Finally, the scavenging rate and reducing power of DMW chitosan and sulfated derivatives increased with their increasing concentration. Moreover, change of DMW sulfated chitosans was the most pronounced within the experimental concentration. However, chelating effect of DMW chitosans were not concentration dependent except for CTS4 and CTS1.