Surface modification of chitosan films. Effects of hydrophobicity on protein adsorption

The surface of chitosan films was modified using acid chloride and acid anhydrides. Chemical composition at the film surface was analyzed by attenuated total reflectance Fourier-transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectroscopy (XPS). ATR-FTIR data verified that the substitution took place at the amino groups of chitosan, thus forming amide linkages, and the modification proceeded to the depth at least 1 microm. Choices of molecules substituted at the amino groups of the glucosamine units did affect the hydrophobicity of the film surface, as indicated by air-water contact angle analysis. The surface became more hydrophobic than that of non-modified film when a stearoyl group (C(17)H(35)CO-) was attached to the films. The reaction of chitosan films with succinic anhydride or phthalic anhydride, however, produced more hydrophilic films. Selected modified films were subjected to protein adsorption study. The amount of protein adsorbed, determined by bicinchoninic acid (BCA) assay, related to the types of attached molecules. The improved surface hydrophobicity affected by the stearoyl groups promoted protein adsorption. In contrast, selective adsorption behavior was observed in the case of the chitosan films modified with anhydride derivatives. Lysozyme adsorption was enhanced by H-bonding and charge attraction with the hydrophilic surface. While the amount of albumin adsorbed was decreased possibly due to negative charges that gave rise to repulsion between the modified surface and albumin. This study has demonstrated that it is conceivable to fine-tune surface properties which influence its response to bio-macromolecules by heterogeneous chemical modification.     

Quaternization of N-aryl chitosan derivatives: synthesis, characterization, and antibacterial activity

Chemical modification of chitosan by introducing quaternary ammonium moieties into the polymer backbone renders excellent antimicrobial activity to the adducts. In the present study, we have synthesized 17 derivatives of chitosan consisting of a variety of N-aryl substituents bearing either electron-donating or electron-withdrawing groups. Selective N-arylation of chitosan was performed via Schiff bases formed by the reaction between the 2-amino groups of the glucosamine residue of chitosan with aromatic aldehydes under acidic conditions, followed by reduction of the Schiff base intermediates with sodium cyanoborohydride. Each of the derivatives was further quaternized using N-(3-chloro-2-hydroxypropyl)trimethylammonium chloride (Quat-188) as the quaternizing agent that reacted with either the primary amino or hydroxyl groups of the glucosamine residue of chitosan. The resulting quaternized materials were water soluble at neutral pH. Minimum inhibitory concentration (MIC) antimicrobial studies of these materials were carried out on Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive) bacteria in order to explore the impact of the extent of N-substitution (ES) on their biological activities. At ES less than 10%, the presence of the hydrophobic substituent, such as benzyl and thiophenylmethyl, yielded derivatives with lower MIC values than chitosan Quat-188. Derivatives with higher ES exhibited reduced antibacterial activity due to low quaternary ammonium moiety content. At the same degree of quaternization, all quaternized N-aryl chitosan derivatives bearing either electron-donating or electron-withdrawing substituents did not contribute antibacterial activity relative to chitosan Quat-188. Neither the functional group nor its orientation impacted the MIC values significantly.     

Synthesis and characterization of N-aryl chitosan derivatives

Selective N-arylation of chitosan was performed via a Schiff bases formed by the reaction between the 2-amino group of glucosamine residue of chitosan with an aromatic aldehyde under acidic condition followed by reduction of the Schiff base intermediate with sodium cyanoborohydride (Borch reduction). Aromatic aldehydes bearing either an electron donating or electron withdrawing substituent were used. The chemical structures and thermal properties of the N-aryl chitosans were characterized by FT-IR, (1)H NMR, (13)C NMR, TGA, and DSC. The extent of N-substitution (ES) was influenced by the molar ratio of the aldehyde to the glucosamine residue of chitosan, the reaction time and the substituent on the aromatic ring. Lower ESs resulted from N-arylation using an aldehyde with an electron donating substituent. A linear relationship between the targeted ES and the ES obtained was observed when aldehydes bearing electron withdrawing substituents were employed.     

Synthesis of methylated chitosan containing aromatic moieties: Chemoselectivity and effect on molecular weight

N-Arylated chitosans were synthesized via Schiff bases formed by the reaction between the primary amino group of chitosan with aromatic aldehydes followed by reduction of the Schiff base intermediates with sodium cyanoborohydride. Treatment of chitosan containing N,N-dimethylaminobenzyl and N-pyridylmethyl substituents with iodomethane under basic conditions led to quaternized N-(4-N,N-dimethylaminobenzyl) chitosan and quaternized N-(4-pyridylmethyl) chitosan. Methylation occurred at either N,N-dimethylaminobenzyl and N-pyridylmethyl groups before the residual primary amino groups of chitosan GlcN units were substituted. The total degree of quaternization of each chitosan varied depending on the extent of N-substitution (ES) and the sodium hydroxide concentration used in methylation. Increasing ES increased the total degree of quaternization but reduced attack at the GlcN units. N,N-dimethylation and N-methylation at the primary amino group of chitosan decreased at higher ES’s. Higher total degrees of quaternization and degrees of O-methylation resulted when higher concentrations of sodium hydroxide were used. The molecular weight of chitosan before and after methylation was determined by gel permeation chromatography under mild acidic condition. The methylation of the N,N-dimethylaminobenzyl derivative with iodomethane was accompanied by numerous backbone cleavages and a concomitant reduction in the molecular weight of the methylated product was observed. The antibacterial activity of water-soluble methylated chitosan derivatives was determined using Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive) bacteria; minimum inhibitory concentrations (MIC) of these derivatives ranged from 32 to 128 μg/mL. The presence of the N,N-dimethylaminobenzyl and N-pyridylmethyl substituents on chitosan backbone after methylation did not enhance the antibacterial activity against S. aureus. However, N-(4-N,N-dimethylaminobenzyl) chitosan with degree of quaternization at the aromatic substituent and the primary amino group of chitosan of 17% and 16–30%, respectively, exhibited a slightly increased antibacterial activity against E. coli.     

Observations of immature and adult stages of the myrmecophilous cetoniine beetle Campsiura nigripennis (Coleoptera: Scarabaeidae)

Several cetoniine species are known or speculated to be associated with ants, based on their specialized morphological characters. However, there are only a few species where biological information on the larval and adult stages is available. Field observations revealed that Campsiura nigripennis spends the immature stages inside elephant dung, and that adult females fly to elephant dung for oviposition. In addition, adult beetles of C. nigripennis intruded into arboreal nests of Oecophylla smaragdina. Specialized morphological characters appear to allow them to tolerate attacks from the ants. Furthermore, the distribution of the beetle in continental Asia largely overlaps that of the Asian elephant, indicating that dung of elephants, in conjunction with that of other large mammals, is fundamental to the biology of C. nigripennis.