A physicochemical, structural, microbiological and kinetic study of hen egg white lysozyme in complexes with alginate and chitosan

Abstract

Complexes of hen egg white lysozyme (HEWL) with alginate (ALG) and chitosan (CS) (natural biodegradable polysaccharides) are important as antimicrobial agents in oral, intranasal and buccal administration. The HEWL–ALG, HEWL–ALG–Ca, [HEWL–ALG]–CS, and [HEWL–ALG–Ca]–CS complexes have been investigated using a combination of physicochemical methods (fluorescent spectroscopy, circular dichroism, microscopy, turbidimetry, microbiology, in vitro cytotoxicity assay). The complexes obtained were non-toxic negatively charged microparticles with high HEWL entrapment efficiency. The [HEWL–ALG]–CS and [HEWL–ALG–Ca]–CS complexes were found to have the altered tertiary structure of HEWL (and/or changes in the polarity of the microenvironment of Trp residues in HEWL molecules). The HEWL entrapment in all studied complexes was carried out with retention of the HEWL activity. Under the long–term storage conditions (1.5 years at 4 °C), Ca++ cations and CS stabilized HEWL. Under physiological conditions, the [HEWL–ALG]–CS and [HEWL–ALG–Ca]–CS complexes demonstrated sustained release of HEWL and resistance to protease action. The [HEWL–ALG]–CS and [HEWL–ALG–Ca]–CS complexes were highly active against Gram-negative Escherichia coli cells. The data obtained are important in the development of antibacterial drugs based on lytic enzymes.