2.4. In Vitro PK/PD Model

A one-compartment in vitro PK/PD model (PASS-402W System, Japan) was employed to evaluate the bactericidal effects of colistin alone or in combination with fosfomycin over a period of 24 h. The apparatus consisted of one central chamber with a capacity of 200 mL that was connected to a reservoir containing CAMHB broth. The temperature of broth within the system was maintained at 37°C. Fresh broth was pumped into the central chamber through a peristaltic pump at a defined flow rate. There was mixing to ensure distribution. Low, medium, and high C_max values of colistin at 0.5, 2, and 5 mg/L as well as C_max of fosfomycin at 100 mg/L were simulated in this PK/PD model. Simulated half-life for colistin and fosfomycin was 4 h and 2.7 h, respectively, as suggested in previously study [18]. Colistin concentrations were modified from the PK profiles in previously published report of in vitro PK/PD study, which maintained a constant concentration of colistin at 0.5, 2, or 5 mg/L [19]. Fosfomycin regimen was also simulated according to the PK parameters achieved in clinic [20]. At the beginning of the experiments, stock solutions of colistin and fosfomycin were injected into the model to achieve the target maximum concentration, and bolus injections were given every 8 h till 24 h. In addition, 25 mg/L of G6P was maintained in the MHB medium throughout the entire experiments. Figure 1 illustrates the targeted steady-state concentration-time profiles of colistin alone or in combination with fosfomycin. In this PK/PD model, synergy is defined as a decrease of ⩾2 log⁡10 in the number of CFU/mL between the combination and its most active component; additivity is defined as a decrease of 1.0 to <2 log⁡10 in the number of CFU/mL between the combination and its most active component [21].

Targeted steady-state concentration-time profiles for colistin (Col, every 8 h) alone or in combination with fosfomycin (Fos, every 8 h). (a) Col 0.5 mg/L, (b) Col 2 mg/L, (c) Col 5 mg/L, (d) Fos 100 mg/L, (e) Col 0.5 mg/L plus Fos 100 mg/L, (f) Col 2 mg/L plus Fos 100 mg/L, and (g) Col 5 mg/L plus Fos 100 mg/L.