Study On Long-circulating Liposomal Daptomycin Against Methicillin-resistant Staphylococcus Aureus

In the face of escalating problems with pathogen control, the development of proper formulations of existing antibiotics is as important as the development of novel antibiotics. Daptomycin is a lipopeptide antibiotic with potent activity against Gram-positive bacteria. Currently, only injectable solution of daptomycin have been approved for clinical use. In the present study, the formulation of PEGylated liposomal daptomycin (PLD) was prepared and optimized, and its efficacy against methicillin-resistant Staphylococcus aureus MRSA252strains was investigated. The obtained PLD had a mean vesicle diameter of (111.5±15.4) nm and a mean percent drug loading of (5.81±0.19)%, with high storage stability. Potent activity of PLD against MRSA was demonstrated in vitro with a more sustained effect than conventional liposomal daptomycin and daptomycin solution. In addition, intravenous administration of a single dose (equal to human use) of PLD significantly increased the survival mice in a MRSA systemic infection model compared with other formulations. Drug distribution in the lung was significantly enhanced following administration of PLD, and no measurable tissue lesions or pathological changes were detected during PLD treatment. Taken together, PEGylated liposomes loaded with daptomycin may represent a promising approach to reduce MRSA infections, especially those involving bloodstream dissemination, such as hematogenous pulmonary infection. The main research topics are following:(1) Establishing a long-circulating liposomes daptomycin analysis methods Determine daptomycin maximum absorption wavelength of221nm, and also identified by HPLC daptomycin chromatographic conditions: Column:Inertsil ODS-SP C18column (150mm×5mm,5μm) Mobile phase:acetonitrile-0.5%ammonium dihydrogen phosphate buffer=40:60Flow rate:0.7mL/min; detection wavelength:221nm Injection volume:10μL; column temperature:30℃Peak time:6.5±0.2min The limit of detection:0.60ng Through recovery experiments, experimental results of this detection method precision accurate and reproducible.(2) Preparation and characterization of long-circulating liposomal daptomycinSeveral factors affect the preparation of daptomycin by examining long-circulating liposomes:the molar ratio of hydrogenated lecithin (HSPC) and cholesterol, fat mass ratio and drug high pressure homogenizer cycles. Eventually get the optimal prescription daptomycin preparation for long-circulating liposomes, hydrogenated lecithin:cholesterol:mPEG2000-DSPE=18:6:1(molar ratio), lipid drug ratio=15:1(mass ratio) hydration fluid pressure homogenizer1000bars after10cycles, you can get more stable long-circulating liposomes daptomycin. The optimal preparation of the obtained liposome particle size (111.5±15.4) nm, Zeta potential (-15.1±2.8) mV, drug loading was (5.81±0.19)%, encapsulation efficiency (92.56±3.15)%, and stable, stored in a refrigerator at4℃almost no leakage after two months, no precipitation occurred.(3) In vitro pharmacodynamic studyMainly study on the growth of MRSA252followed influenced by three formulations (DS, CLD, PLD), results showed that, PLD inhibition of MRSA252more lasting effect.(4) Pharmacokinetic and biodistribution studyThe results of surface long-circulating liposomes daptomycin extended daptomycin in rat blood circulation time, and significantly increased the amount of daptomycin distribution in the liver, spleen and lungs.(5) In vivo pharmacodynamic study and safety evaluation of long-circulating liposomal daptomycinBy comparing the DS, CLD, PLD for MRSA252infected mice induced systemic therapeutic effect compared the results showed that daptomycin intravenously administered once a long-circulating liposomes can make the survival of infected mice increased to80%. Safety evaluation results showed that daptomycin long-circulating liposomes had no obvious toxicity in mice.