Preparation of biguanidine quaternary ammonium salts grafted chitosan with enhanced antibacterial and antibiofilm activities.

N-(4-N'-pyridine-benzylcarbonyl chloride) chitosan (CBPyC), N-p-biguanidine benzoyl chitosan (CSBG), and N-(p-biguanidine -1-pyridine-4-benzylcarbonyl chloride) chitosan (CSQPG) were synthesized. The structures of prepared chitosan derivatives were characterized using nuclear magnetic resonance spectroscopy (NMR) and ultraviolet-visible (UV-vis) spectroscopy, and the degree of substitution was determined through elemental analysis (EA) and evaluated on the basis of the integral values in 1H NMR. The antibacterial activities of chitosan derivatives against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were investigated in vitro using antibacterial rate, minimal inhibitory concentration and minimum bacterial concentration assays. The antibiofilm activity was also assessed using the crystal violet assay. CSQPC exhibited higher antibacterial and antibiofilm activities against E. coli and S. aureus compared to CBPyC and CSBG. The antibacterial rate of CSQPG against E. coli and S. aureus at a concentration of 0.5 mg/mL was 43.3% and 100%, respectively. The biofilm inhibition rate of CSQPG at 0.5 MIC against E. coli and S. aureus was 56.5% and 69.1%, respectively. At a concentration of 2.5 mg/mL, the biofilm removal rates of E. coli and S. aureus were 72.9% and 90.1%, respectively. The antibacterial and antibiofilm activities of CSQPG were better than CSBG and CBPyC, and the combination of guanidine and quaternary ammonium further improved the positive charge density of chitosan and enhanced its antibacterial activity.

Effect of the structure of chitosan quaternary phosphonium salt and chitosan quaternary ammonium salt on the antibacterial and antibiofilm activity.

N-(4-N', N', N'-trimethylphosphonium chloride) benzoyl chitosan (TMPCS), N-(4-N', N', N'-triphenylphosphonium chloride) benzoyl chitosan (TPPCS), and N-(4-N', N', N'-trimethylmethanaminium chloride) benzoyl chitosan (TMACS) were synthesized. The structures of the products were characterized by Fourier transform infrared spectroscopy, Nuclear magnetic resonance spectroscopy and ultraviolet-visible spectroscopy. Their antibacterial activities against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were investigated in vitro using the antibacterial rate, minimal inhibitory concentration (MIC) and minimum bactericidal concentration (MBC), the antibiofilm activity was investigated by crystal violet assay. The antibacterial assessment revealed that the chitosan quaternary phosphonium salts of similar structure had superior antibacterial activity than chitosan quaternary ammonium salt. The antibacterial rate of CS, TMPCS, TPPCS and TMACS against E. coli at 0.5 mg/mL was 10.4 %, 42.0 %, 58.5 % and 21.6 % respectively. At the same concentration, the antibacterial rate of TMPCS, TPPCS and TMACS against S.aureus was all up to 100 %. The biofilm inhibition rate of CS, TMPCS, TPPCS and TMACS at a half of MIC against E.coli was 28.4 %, 33.9 %, 56.6 % and 57.6 % respectively, and against S.aureus was 30.8 %, 53.8 %, 62.2 % and 58.5 % respectively. The biofilm removal rate of CS, TMPCS, TPPCS, TMACS against E.coli at 2.5 mg/mL was 20.6 %, 46.4 %, 48.9 % and 41.6 % respectively, and against S.aureus at 2.5 mg/mL was 41.5 %, 60.4 %, 69.9 % and 59.01 % respectively.

Synthesis, characterization, and antibacterial activity of N,O-quaternary ammonium chitosan.

N,N,N-Trimethyl O-(2-hydroxy-3-trimethylammonium propyl) chitosans (TMHTMAPC) with different degrees of O-substitution were synthesized by reacting O-methyl-free N,N,N-trimethyl chitosan (TMC) with 3-chloro-2-hydroxy-propyl trimethyl ammonium chloride (CHPTMAC). The products were characterized by (1)H NMR, FTIR and TGA, and investigated for antibacterial activity against Staphylococcus aureus and Escherichia coli under weakly acidic (pH 5.5) and weakly basic (pH 7.2) conditions. TMHTMAPC exhibited enhanced antibacterial activity compared with TMC, and the activity of TMHTMAPC increased with an increase in the degree of substitution. Divalent cations (Ba(2+) and Ca(2+)) strongly reduced the antibacterial activity of chitosan, O-carboxymethyl chitosan and N,N,N-trimethyl-O-carboxymethyl chitosan, but the repression on the antibacterial activity of TMC and TMHTMAPC was weaker. This indicates that the free amino group on chitosan backbone is the main functional group interacting with divalent cations. The existence of 100 mM Na(+) slightly reduced the antibacterial activity of both chitosan and its derivatives.

Relationship between N,N-dialkyl chitosan monolayer and corresponding vesicle.

The properties of N,N-dialkyl chitosan monolayers and corresponding vesicles have been studied by LB technique and drug-release experiment. With increasing molecular weight of chitosan backbone and/or length of alkyl chain, both the compressibility and collapse pressure of N,N-dialkyl chitosan monolayer are enhanced. The experiments on drug-release behavior of N,N-dialkyl chitosan vesicles show that the drug-release rate and the equilibrium drug-release ratio are decreased with increasing the compressibility of corresponding monolayer. It is worth noticing that there is a linear relationship between the compressibility of N,N-dialkyl chitosan monolayers and the equilibrium drug-release ratio of the vesicles.