黄连提取物对金黄色葡萄球菌的抑菌活性及作用机理

doi:10.13304/j.nykjdb.2024.0132

摘要/Abstract

摘要：

为了解黄连提取物对金黄色葡萄球菌的抑菌机理，通过生长曲线、氧化损伤试验、细胞壁膜分析、蛋白质分析和DNA分析，探讨黄连提取物对金黄色葡萄球菌的作用机制。结果表明，黄连提取物对金黄色葡萄球菌有明显的抑制效果，最小抑菌浓度为2.60 mg·mL^-1。黄连提取物可引起金黄色葡萄球菌胞内活性氧水平增加506.90%，胞外β-半乳糖苷酶活性和核酸含量分别增加258.28%和23.82%，碘化丙啶荧光染色也明显多于对照；总蛋白合成量、细胞代谢活力以及琥珀脱氢酶活性分别降低30.83%、30.89%、54.05%。基因组DNA的紫外吸收光谱和竞争性荧光光谱显示，黄连提取物可以与金黄色葡萄球菌DNA进行小沟结合或静电结合。以上结果表明，黄连提取物主要通过增加胞内活性氧水平，破坏细胞壁膜完整性及通透性来达到抑菌效果。研究结果为黄连的综合开发利用及其在饲料添加剂领域的应用提供借鉴。

关键词: 金黄色葡萄球菌, 黄连提取物, 抑菌机理

Abstract:

In order to understand the bacteriostatic mechanism of rhizoma coptidis extract （RCE） on Staphylococcus aureus， the mechanism of action of RCE on Staphylococcus aureus was explored through growth curve， oxidative damage experiment， cell wall membrane analysis， protein analysis and DNA analysis. The results showed that RCE had a significant inhibitory effect on Staphylococcus aureus with a minimum inhibitory concentration of 2.60 mg·mL^-1. RCE induced an increase in intracellular reactive oxygen species level of Staphylococcus aureus by 506.90%， an increase in extracellular β-galactosidase activity and nucleic acids by 258.28% and 23.82%， respectively， and also a significant amount of propidium iodide fluorescent staining than that of the control； the amount of total protein synthesis， cellular metabolic vigor and succinic dehydrogenase activity decreased by 30.83%， 30.89% and 54.05%， respectively. Ultraviolet absorption spectra and competitive fluorescence spectra of genomic DNA suggested that the RCE could be either groove-bound or electrostatically bound to Staphylococcus aureus DNA. As the above shows， RCE mainly achieved its bacteriostatic effect by increasing the level of intracellular reactive oxygen species and destroying the integrity and permeability of the cell wall membrane. Above results provided references for the comprehensive development and utilization of rhizoma coptidis and its application in the field of feed supplement.

Key words: Staphylococcus aureus, rhizoma coptidis extract, bacteriostatic mechanism

中图分类号:

S816.7

周瑾, 梁海运, 孙佳慧, 张舒涵, 宋丽雅. 黄连提取物对金黄色葡萄球菌的抑菌活性及作用机理[J]. 中国农业科技导报, 2025, 27(3): 143-152.

Jin ZHOU, Haiyun LIANG, Jiahui SUN, Shuhan ZHANG, Liya SONG. Bacteriostatic Activity and Mechanism of Action of Rhizoma Coptidis Extracts on Staphylococcus aureus[J]. Journal of Agricultural Science and Technology, 2025, 27(3): 143-152.

图/表 9

图1 RCE对金黄色葡萄球菌作用后的生长曲线

Fig. 1 Kinetic growth profile of Staphylococcus aureus upon exposure to RCE

图2 RCE对金黄色葡萄球菌作用后的荧光强度注：****表示在P<0.000 1水平差异显著。

Fig. 2 Fluorescence intebsity of Staphylococcus aureus after exposure to RCENote：**** indicates significant differences at P<0.000 1 level.

图3 PI标记结果A：对照组明场结果；B：RCE作用后的明场结果；C：对照组暗场结果；D：RCE作用后的暗场结果

Fig. 3 Result of PI-labeling experimentA： Bright field result of control group； B： Bright field result after RCE action； C： Dark field result of the control group； D： Dark field result after RCE

图4 RCE作用于金黄色葡萄球菌后胞外β-半乳糖苷酶含量注：**表示差异在P<0.01水平显著。

Fig. 4 Content of β-galactosidase in Staphylococcus aureus after exposure to RCENote：** indicates significant difference at P<0.01 level.

图5 RCE作用于金黄色葡萄球菌后的核酸泄漏量注：*和**分别表示两组间在P<0.05和P<0.01水平差异显著。

Fig. 5 Amount of nucleic acid leakage from Staphylococcus aureus after exposure to RCENote：* and ** indicate significant differences between two groups at P<0.05 and P<0.01 levels，respectively.

图6 RCE作用于金黄色葡萄球菌后细胞内总蛋白含量注：***表示差异在P<0.001水平显著。

Fig. 6 Intracellular total protein content of Staphylococcus aureus after exposure to RCENote：*** indicates significant difference at P<0.001 level.

图7 RCE作用于金黄色葡萄球菌后细胞的代谢活力和酶活力A：细胞活力；B：细胞总ATPase活性；C：细胞SDH活性。*、**和****分别表示在P<0.05、P<0.01和P<0.000 1水平差异显著；ns表示差异不显著

Fig. 7 Activity of cellular metabolism and enzyme of Staphylococcus aureus after exposure to RCEA： Cell viability； B： Cellular total ATPase activity； C： Cellular SDH activity. *，**and **** indicate significant differences at P<0.05，P<0.01 and P<0.000 1 levels，respectiveiy；ns indicates not significant

图8 RCE与金黄色葡萄球菌DNA紫外光谱结果

Fig. 8 Ultraviolet （UV） spectroscopy results of the interaction between RCE and Staphylococcus aureus DNA

图 9 RCE与DAPI/EB竞争性结合DNA的荧光光谱图A：DAPI-DNA体系；B：EB-DNA体系

Fig. 9 Fluorescence emission spectra of DAPI/EB-DNA in the presence of RCEA：DAPI-DNA system； B：EB-DNA system

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