Inhalable Levofloxacin Liposomes Complemented with Lysozyme for Treatment of Pulmonary Infection in Rats: Effective Antimicrobial and Antibiofilm Strategy

Treatment of bacterial infections becomes increasingly complicated due to increasing bacterial resistance and difficulty in developing new antimicrobial agents. Emphasis should be laid on improvising the existing treatment modalities. We studied the improved antimicrobial and antibiofilm activity of levofloxacin (LFX) and lysozyme (LYS) in microbiological studies. LFX at sub-minimum inhibitory concentration with LYS eradicated > 85% of preformed biofilm. LFX was actively loaded into the liposomes using pH gradient method and was spray-dried with LYS solution. Percent entrapment of LFX in liposome was > 80% and prolonged cumulative release of 85% LFX at the end of 12 h. In vitro lung deposition study and solid-state characterization for spray dried LFX liposome in combination with LYS (LFX liposome-LYS) was performed. Co-spray dried product had mass median aerodynamic diameter ranging < 5 mu m. In pharmacodynamic study, Staphylococcus aureus infected rats were treated with LFX liposome-LYS. Lungs, bronchoalveolar lavage fluid (BALF), and nasal fluid were evaluated for microbial burden. Expression of cytokine levels in BALF and serum were also studied by ELISA. In addition, mRNA expression for lung inflammatory mediators and lung myeloperoxidase activity were carried out. Further, lungs and histological changes were observed grossly. Untreated infected rat lungs demonstrated higher mRNA expression for inflammatory markers, cytokine levels, and microbial load compared to vehicle control. Conversely, LFX liposome-LYS significantly abated these adverse repercussions. Histology findings were also in agreement of above. Acute toxicity study revealed safeness of LFX liposome-LYS. Our findings confirm LFX liposome-LYS exhibited prolonged, improved antibiofilm and antimicrobial efficacy in treating S. aureus infection.