为规范抗菌肽抑制丝状真菌的活力评价方法,采用多粘菌素B为抗菌肽研究对象,以黑曲霉(Aspergillus niger ATCC 16404)和产黄青霉(Pencillinm chrysogenum ATCC 10106)为指示菌,选用察氏培养基(20 mL培养基/90 mm平板)比较孢子萌发抑制法和菌丝生长抑制法,获得适合丝状真菌的测定方法,并进一步研究该方法的最适测定条件。结果表明,菌丝生长抑制法更适用于多粘菌素B抑制2株丝状真菌的定量检测。其中多粘菌素B抑制Aspergillus niger ATCC 16404的最适条件:指示菌培养48 h,接种菌饼直径(8.00±0.02) mm,在此条件下,测得多粘菌素B的EC50为0.68 mg/mL;多粘菌素B抑制Pencillinm chrysogenum ATCC 10106的适宜条件:指示菌培养72 h,接种菌饼直径(5.80±0.02) mm,此时多粘菌素B的EC50为0.45 mg/mL。本研究建立的抗菌肽抑制丝状真菌的活力测定方法具有普遍适用性,可为抗菌肽活性测定方法的规范及评价标准的建立奠定基础。
In order to standardize the evaluation methods of antimicrobial peptides against filamentous fungi,polymyxin B was selected as the antifungal peptide,Aspergillus niger ATCC 16404 and Pencillinm chrysogenum ATCC 10106 were chosen as the bioindicators of fungi to determine the measure method in this study. The evaluation methods of polymyxin B against filamentous fungi were determined on czapek’s medium(20 mL medium per 90 mm petri dish)by comparing with two common methods of conidial germination inhibition and mycelial growth inhibition. The suitable determination conditions were then further optimized. The results showed that the evaluation method of mycelial growth inhibition was more suitable for the quantitative determination of polymyxin B against filamentous fungi. The optimal test conditions for polymyxin B against A. niger ATCC 16404 were as follows:The culture time of fungal pathogen was 48 h,the diameter of mycelium plug was(8.00±0.02)mm,and the EC50 of polymyxin B was 0.68 mg/mL under these conditions. The suitable conditions for polymyxin B against P. chrysogenum ATCC 10106 were as follows:The culture time was 72 h and the mycelium plug diameter was(5.80±0.02)mm,and the EC50 of polymyxin B was 0.45 mg/mL. The determination method of antifungal activity for antimicrobial peptides against filamentous fungi had a widely applicability,and it could provide a new foundation for the standard and establishment of determination method on the antimicrobial peptide against filamentous fungi.