Preparation Of Antifungal Peptide Aps, Biological Activity And Its Mechanism Of Anti-fungal

The incidental of fungal infections has increased and represent a new challenging problem for clinicians because of the increase in the number of people whose immune system are compromised by AIDS, cancer therapy and organ transplantation and the abuse of amtibiotic. The toxicity of the current synthesized antibiotics and the development of resistant fungal strains will cause the already limited antifungal drug become more shortage. It is therefore imperative to find antifungal compounds that are effective, broad-spectrum and safe while using antifungal medicine in reason.Recently, antimicrobial peptides with their fundamental role in host innate immunity defense mechanism have received increasing attention because of their potent antifungal activity and unique antifungal mechanism. APS, found by Prof. Pei Yan, et al, is a novel antifungal peptide isolated from Bacillus cereus S-1. It has been reported that APS has potent antifungal activity against various phytopathogenic fungi. This paper optimizes the process of isolation and purification, detects cytotoxicity of APS, observes curative effect on cavies whose skin has infected by Candida albicans, so that we can primarily estimate their antifungal therapeutic probability and it also elucidats possible antifungal mechanism of APS.On the base of having been purified by silica gel previously, the process of combining subsection elution and linear gradient elution with Rsource RPC and RPC Sephasil Peptide C18 column was established. And a class of functional components were obtained, named as APS1, APS2, APS3, APS4 and APS5. The cytotoxicity of APS on L-02, ECV-304, 293 and 3T3 cell was investigated with the observation of the cell morphology and MTT assay.APS5 has the highest cytotoxicity and the APS1 has the lowest cytotoxicity. Parts of APS have lower cytotoxicity than amphotericin B. The hemolytic activities of APS on human erythrocytes were measured by hemolytic assay. The hemolytic activity of APS 1-3 are lower than amphotericin B while APS4-5 are almost the same as amphotericin B. The animal experiment indicats that curative effect of APSl is similar with fluconazole. The difference between the high dose and the low dose was the time at which the curative effect had presented. The research of the antifungal mechanism showed that APSl could not only inhibited the growth of Saccharomyces cerevisiae but also killed them rapidly. After incubated with APSl for 2.5h, Saccharomyces cerevisia had been dyed by PL Transmission electronic microscope observation showed that the membrane and cell wall of the treated cells with APSl changed greatly. The results above indicate that APS 1 may killed the treated cells by changing the membrane permeability.The optimized purification process of APS was established. We found that APS has little cytotoxicity, some of their cytotoxicity are lower than amphotericin B, and APS has curative effect on the cavy whose skin has infected by Candida albicans. Base on the study results above, APS shows latency value for becoming a novel antifungal drug in future. The mechanism of its action is most likely to cause membrane permeation and the death of fungi.