Study On Amphotericin B-loaded Micelles

Fungal infections have become a health concern due to their severe morbidity and mortality.Amphotericin B(Ammphotericin B,AmB)is a clinically effective antifungal drug.Its monomers and aggregates can interact with ergo sterol on the fungal cell membrane,unfortunately,its aggregates can non-selectively destroy mammalian cells.Its clinical applicationits was also limited owing to poor water solubility,toxic and side effects such as nephrotoxicity,cardiotoxicity,and liver toxicity.Amphiphilic polymer materials usually consist of a hydrophobic segment and a hydrophilic segment,which can be used as a drug carrier to encapsulate a poorly soluble drug in a hydrophobic core.Drug-loaded micelles have the advantages of good stability,increased solubility of poorly soluble drugs,reduced adverse reactions,and improved drug bioavailability.They are an excellent drug-loaded system that has been newly developed in recent years.In this paper,the target product monomethoxy poly(ethylene glycol)-g-PEI-g-ALA(MPEG-PEI-ALA)was synthesized by using α-linolenic acid,poly(ethylene imine),and monomethoxy polyethylene glycol obtained as starting materials.The structure of the target product was identified by nuclear magnetic resonance spectroscopy(1H-NMR)and infrared spectroscopy(IR).The synthesized product’s critical micelle concentration and hemolysis activity were performed.The dialysis method was used to explore the preparation process of AmB drug-loaded micelles(AmB-M),and the prepared micelles were studied by X-ray diffraction,IR,Zeta potential,morphology,stability,aggregation state,hemolysis activity and in vitro release.Using Candida albicans as a fungal model,the drug-loaded micelles were performed with tests as follows:in vitro antifungal activity,diameters of inhibition zone,biofilm inhibition activity,DAPI staining apoptotic phenotype,in vivo antifungal activity,and in vivo renal toxicity.The results showed that the optimal process for preparing AmB-M was:the molar ratio ofα-linolenic acid to MPEG-PEI was 2:1,the ratio of the amount of carrier to the amount of drug was 10:1,and the ultrasound time was 60 min.The optimal drug loading and encapsulation efficiency of the prepared drug-loaded micelles were 8.87±0.13%and 87.35±1.06%,respectively.The average particle size was 229.05±3.13 nm and the zeta potential was 15.80±0.12 mV.It had good storage stability within 21 days,its hemolytic properties meet the requirements of biocompatibility,and the degree of aggregation was lower than the commercially available injection(AmB-I).Both the IR and X-ray diffraction patterns showed that amphotericin B was successfully encapsulated into the micelles.After encapsulated into micelles,crystalline state of the drug changed into the amorphous state.In vitro release experiments showed that the release rate of AmB-M was slower than that of the drug from AmB-I,and AmB-M had abetter sustained release effectIn vitro experiments showed that,using Candida albicans as a model,the MIC90 value of AmB-I was twice that of AmB-M.The inhibition zone experiment was performed using the Oxford Cup method,and a visible inhibiton zone was observed.The AmB-I and AmB-M had similar diameters of inhibiton zone.XTT reduction-menaquinone experiments showed that at a concentration of 0.5 μg/mL AmB,AmB-M and AmB-I have moderate and equivalent levels of in vitro biofilm inhibition activity.DAPI staining fluorescence microscopy studies showed that AmB-M could cause Candida albicans apoptosis to the same extent as AmB-IIn vivo experiments had shown that AmB-M demontrated less renal toxicity than AmB-I A mouse fungal infection model was established with Candida albicans intraperitoneal infection.The results showed that the in vivo activity of AmB-M was similar to AmB-I.The organ toxicity in the AmB-M treatment group was lower than that for the AmB-I treatment group.