Preparation and characterization of chitosan microspheres for doxycycline delivery

Doxycycline-loaded chitosan microspheres were developed using a novel water-in-oil emulsion technique, involving oil phase ionic gelation. Microspheres were prepared by using 6% v/v of chitosan (3% w/v in acetic acid), soya oil–n-octanol oil mixture (1:2 v/v) as continuous phase and 5% span 80 as emulsifier. Doxycycline was entrapped by equilibrium swelling method with 8.4% total entrapment. The drug-loaded spheres were spherical with smooth surface morphology. The MTT assay showed that doxycycline-loaded microspheres were able to improve the percentage cell viability in comparison to the pure drug. In vitro release studies showed that a burst release of 42% in 6 h was achieved and maintained an equilibrium concentration of 72% in 24 h. Assessment of antibacterial activity showed that doxycycline was able to exhibit a minimum microbicidal concentration (MIC) of 16.5, 17.4, 11.2 and 98.3 μg against Klebsiella pneumoniae (ATCC 15380), Escherichia coli (ATCC 25922), Staphylococcus aureus (ATCC 9144) and Pseudomonas aeruginosa (ATCC 25619), respectively. Gelatin zymography studies revealed that it could inhibit MMP 2 and MMP 9 at sub-antimicrobial concentration. The present investigation provides scope for using doxycycline-loaded chitosan microspheres for healing infected wounds.