牛蛙背皮抗菌肽粗提物的抑菌活性

Bacteriostatic activity of antimicrobial peptide crude extract from bullfrog skin of back

研究旨在分析牛蛙背皮抗菌肽粗提物的抑菌活性.通过甲醇浸提法制备了牛蛙背部皮肤的抗菌肽粗提物,以琼脂糖扩散法检测粗提物的抑菌活性,用倍比稀释法测定粗提物的最小抑菌浓度(MIC),通过测定生长曲线和杀菌动力学曲线分析粗提物的抑菌特性,借助显微镜观察菌体形态和染色变化,用pH计测定细菌培养基的pH值.结果表明:牛蛙背皮抗菌肽粗提物对大肠杆菌和金黄色葡萄球菌具有抑菌活性,MIC分别为62.5μg/m L和31.25μg/m L,但对白色念珠菌没有抑菌活性;在低质量浓度(1×MIC)时,在迟缓期没有抑菌活性,但从对数期开始发挥抑菌活性;在高质量浓度(3×MIC)时,10 min内即开始发挥抑菌活性;粗提物作用后的大肠杆菌和金黄色葡萄球菌呈现相反的革兰染色结果,菌体形态从杆状或球状变为丝状;培养2 h后,粗提物可提高大肠杆菌培养液的pH值,但对金黄色葡萄球菌培养液的pH值没有影响.结果提示,牛蛙背皮抗菌肽粗提物对大肠杆菌和金黄色葡萄球菌有较好的抗菌活性,为后续研究和临床应用奠定了基础.

The bacteriostatic activity of antimicrobial peptide crude extract from bullfrog skin of back was analyzed in this paper.Antimicrobial peptide crude extract was prepared with methanol extraction.Antibacterial activity of crude extract was detected with agarose diffusion method.Minimal inhibition concentration of crude extract was measured with double dilution method.Antibacterial property of crude extract was assayed with growth curve and killing kinetics of bacterium.Morphological and color changes of bacteria treated with crude extract were observed with microscope.The pH value of bacterial cultural liquid was determined with pH meter.The results showed that antimicrobial peptide crude extract from bullfrog skin of back had antibacterial activity against Escherichia coli and Staphylococcus aureus with the MIC of 62.5 μg/m L and 31.25 μg/m L respectively,but had no activity against Candida albicans.Crude extract with low concentration（1×MIC） had no activity in lag phase,but had activity from logarithmic phase.While crude extract with high concentration（3×MIC） had activity within 10 min.Compared with normal bacteria,the Gram stained results of E. coli and S. aureus treated with crude extract were opposite,and the morphology were changed from rhabditiform or sphericity to filiform.And the pH value of cultural liquid of E. coli treated with crude extract was high than control group,while there were no changes significantly on the pH value of cultural liquid of S. aureus.These findings indicated that antimicrobial peptide crude extract from bullfrog skin of back had bacteriostatic activity against E. coli and S. Aureus,which lay a good foundation for following research and clinical practice.