Influence of a Pulmonary Surfactant on the In Vitro Activity of Tobramycin against Pseudomonas aeruginosa

Abstract

Delivery of therapeutic concentrations of drugs to the lung periphery is crucial in treatment of many infectious and inflammatory diseases of the lung. Conventional drug treatment for serious pulmonary infections consists of systemic administration of drug in order to achieve concentrations in the bloodstream that will reach therapeutic results. Advantages to topical drug administration to the lungs via aerosol are well known (3). However, the amount of drug that can be inhaled as aerosol is frequently small, thereby limiting the amount of drug delivered to the distal portion of the lung (1). Intratracheal instillation of surfactant, in the animal model, increased the fractional area of lung exposed and improved the uniformity of spread of radioactive label in the lung compared to the saline control (2). We examined the effect of physical perturbation with the mixture, antibiotic, and surfactant, via heat and sonication and stability over time as a function of the MIC for Pseudomonas. Tobramycin and surfactant (Exosurf suspension) were combined in cationic adjusted Mueller-Hinton broth (CAMHB) to create mixtures containing 8, 16, 32, or 64 μg of surfactant per ml, all with 4 μg of tobramycin per ml (4). One aliquot of each mixture was heated at 80°C for 30 min, one was sonicated at room temperature for 30 min, and a third was held at room temperature for 30 min without additional treatment. Serial twofold dilutions of each aliquot were prepared in CAMHB in microdilution trays (final volume, 100 μl/well) and inoculated with approximately 5 × 105 CFU of Pseudomonas aeruginosa ATCC 27853. Serial dilutions of tobramycin in CAMHB and separate serial dilutions of surfactant in CAMHB were inoculated with the same organism to serve as controls. All trays were incubated for 18 h at 35°C, and the MIC was determined visually. When tobramycin and surfactant were mixed, sonicated, and heated to 80°C for 30 min, there was no in vitro reduction in MIC for P. aeruginosa. To assess the stability of mixtures of tobramycin and surfactant, we prepared a stock mixture containing 27 mg each of tobramycin and surfactant per ml. Dilutions of this mixture were prepared immediately to give final concentrations of tobramycin ranging from 0.06 to 8 μg/ml in CAMHB. Controls, inoculation, incubation, and examination were identical to those described above. The remaining stock mixture was stored at 4°C for 3 weeks, and the MIC was determined at 1 week and again at 3 weeks. No decline in the MIC of the mixture was found when it was stored in a refrigerated environment for up to 3 weeks. van’t Veen and coworkers (5) found a reduction in bactericidal activity of tobramycin when tobramycin was mixed with surfactant which could be compensated for by increasing the tobramycin concentration. A possible explanation of the difference from our study may be the difference in the surfactants. Exosurf, a synthetic compound, does not contain protein, whereas van’t Veen et al. used a surfactant obtained from the bovine lung containing a protein component.