摘要
目的探讨不同浓度安石榴苷对牙龈卟啉单胞菌、具核梭杆菌在浮游状态和生物膜状态下的抑菌活性的作用。方法(1)配置浓度为1×10^(8)CFU/mL的牙龈卟啉单胞菌、具核梭杆菌悬液,厌氧条件下分别用0.125、0.25、0.5、1、2、4、8 g/L的安石榴苷溶液处理48 h,测定安石榴苷最低抑菌浓度(minimum inhibitory concentration,MIC)及最低杀菌浓度(minimum bactericidal concentration,MBC)。(2)将牙龈卟啉单胞菌、具核梭杆菌悬液分别分为1/2 MIC组(1/2MIC浓度安石榴苷药液)、MIC组(MIC浓度安石榴苷药液)、2 MIC组(2 MIC浓度安石榴苷药液)、阳性对照组(复方氯己定含漱液)和阴性对照组(菌悬液),厌氧条件下培养0、4、8、12、24、48 h时,应用酶标仪测定600 nm波长处光密度(optical density,OD)值。(3)将牙龈卟啉单胞菌、具核梭杆菌悬液分别分为MIC组、1/2 MIC组、1/4 MIC组、1/8 MIC组和阴性对照组,厌氧条件下培养48 h,应用紫外可见分光光度计测定600 nm波长处OD值,计算细菌黏附抑制率。(4)厌氧条件下将2种菌悬液与不同浓度安石榴苷溶液共培养制备生物膜,测定生物膜最低形成抑制浓度;厌氧条件下制备2种菌悬液生物膜,应用不同浓度安石榴苷溶液处理24 h,测定最低清除已形成生物膜浓度。结果(1)安石榴苷对牙龈卟啉单胞菌、具核梭杆菌的MIC均为2 g/L,MBC均为4 g/L。(2)安石榴苷处理4、8、12、24、48 h,1/2 MIC组、MIC组、2 MIC组、阳性对照组和阴性对照组牙龈卟啉单胞菌、具核梭杆菌OD值比较差异均有统计学意义(P<0.05)。随安石榴苷浓度增加,作用时间延长,其对牙龈卟啉单胞菌、具核梭杆菌的抑制作用增强。(3)MIC组、1/2 MIC组、1/4 MIC组牙龈卟啉单胞菌(0.423±0.026、0.764±0.009、1.243±0.023)、具核梭杆菌(0.180±0.007、0.366±0.009、0.612±0.017)细菌黏附OD值均低于1/8 MIC组(1.533±0.015、0.768±0.008)和阴性对照组(1.578±0.058、0.796±0.014)(P<0.05),且依次增高(P<0.05),1/8 MIC组细菌黏附OD值与阴性对照组比较差异无统计学意义(P>0.05)。(4)安石榴苷对牙龈卟啉单胞菌、具核梭杆菌的生物膜最低形成抑制浓度均为4 g/L,最低清除已形成生物膜浓度分别为16、8 g/L。结论安石榴苷是一种天然药物,对牙周主要致病菌的浮游及生物膜状态有一定的抑制作用。
Objective To investigate the antibacterial activities of punicalagin at different concentrations against Porphyromonas gingivalis and Fusobacterium nucleatum in floating state and biofilm state.Methods(1)The suspensions of Porphyromonas gingivalis and Fusobacterium nucleatum with a concentration of 1×10^(8) CFU/mL were prepared and treated with 0.125,0.25,0.5,1,2,4 and 8 g/L of punicalagin solution for 48 h respectively under anaerobic condition.The minimum inhibitory concentration(MIC)and minimum bactericidal concentration(MBC)of punicalagin were detected.(2)The suspensions of Porphyromonas gingivalis and Fusobacterium nucleatum were divided into 1/2MIC group(1/2 MIC concentration of punicalagin),MIC group(MIC concentration of punicalagin),2MIC group(2 MIC concentration of punicalagin),positive control group(compound chlorhexidine gargle)and negative control group(bacterial suspension).After 0,4,8,12,24 and 48 h culture under anaerobic condition,the optical density(OD)value at 600 nm wavelength was measured by enzyme label.(3)The suspensions of Porphyromonas gingivalis and Fusobacterium nucleatum were divided into MIC group,1/2MIC group,1/4MIC group,1/8MIC group and negative control group.The suspensions were cultured for 48hunder anaerobic condition.The OD value at 600nm wavelength was measured by ultraviolet visible spectrophotometry,and the bacterial adherence inhibitory rate was calculated.(4)Biofilm was prepared by co-culture of two bacterial suspensions with different concentrations of punicalagin under anaerobic condition,and minimum biofilm inhibitory concentration was measured.Two kinds of suspension biofilms were prepared under anaerobic condition,and different concentration of punicalagin was applied for 24hto detect the minimum biofilm reduction concentration.Results(1)The MICs and MBCs of punicalagin against Porphyromonas gingivalis and Fusobacterium nucleatum were 2g/L and 4g/L,respectively.(2)There were significant differences in OD values of Porphyromonas gingivalis and Fusobacterium nucleatumin 1/2 MIC group,MIC group,2 MIC group,positive control group and negative control group after 4,8,12,24and 48htreatment with punicalagin(P<0.05).With the increase of concentration of punicalagin and the prolongation of action time,its inhibitory efficacies on Porphyromonas gingivalis and Fusobacterium nucleatum were enhanced.(3)The adherence OD values of Porphyromonas gingivalis and Fusobacterium nucleatumincreased gradually in turn in MIC group(0.423±0.026,0.180±0.007),1/2MIC group(0.764±0.009,0.366±0.009)and 1/4MIC group(1.243±0.023,0.612±0.017)(P<0.05),which were lower than those in 1/8MIC group(1.533±0.015,0.768±0.008)and negative control group(1.578±0.058,0.796±0.014)(P<0.05),and showed no significant differences between 1/8MIC group and negative control group(P>0.05).The adherence inhibitory rates of Porphyromonas gingivalis(73.20%,51.59%,21.25%,2.87%)and Fusobacterium nucleatum(77.39%,54.02%,23.12%,3.52%)decreased gradually in turn in MIC,1/2MIC,1/4MIC and 1/8MIC groups(P<0.05).(4)For Porphyromonas gingivalis and Fusobacterium nucleatum,the minimum biofilm inhibitory concentrations of punicalagin were 4 g/L and 4 g/L,while the minimum biofilm reduction concentrations were 16 g/L and 8 g/L,respectively.Conclusion Punicalagin is a kind of natural drug,with a certain inhibitory efficacy on the main periodontal pathogens in floating state and biofilm state.
作者
苏比努尔·阿尔肯
伊尔凡·努尔买买提
古丽努尔·阿吾提
Subinuer AERKEN;Yierfan NUERMAIMAITI;Gulinuer AWUTI(Department of Periodontal Disease,the First Affiliated Hospital of Xinjiang Medical University,Affiliated Stomatological Hospital of Xinjiang Medical University,Xinjiang Uygur Autonomous Region 830054,China)
出处
《中华实用诊断与治疗杂志》
2023年第3期273-278,共6页
Journal of Chinese Practical Diagnosis and Therapy
基金
中华口腔医学会西部口腔医学临床科研基金(CSA-W2020-11)
新疆维吾尔自治区自然科学基金项目(2011211A071)
新疆维吾尔自治区卫生与健康适宜技术推广项目(SYTG-202115)。
关键词
牙周炎
安石榴苷
牙龈卟啉单胞菌
具核梭杆菌
菌斑生物膜
黏附抑制
生长曲线
抑菌活性
periodontitis
punicalagin
Porphyromonas gingivalis
Fusobacterium nucleatum
plaque biofilm
adherence inhibition
growth curve
antibacterial activity
