Antibacterial Effect of Slightly Acidic Electrolyzed Water on Enterococcus faecalis Biofilm in Root Canal in Vitro
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摘要:
目的 研究微酸性电解水（slightly acidic electrolyzed water，SAEW）作为根管冲洗液对根管内粪肠球菌（Enterococus faecalis，E. faecalis）生物膜的抗菌作用。 方法 离体牙根管内培养E.f生物膜，以1%次氯酸钠（sodium hypochlorite，NaClO）溶液为阳性对照，分别用浓度为50 mg/L、60 mg/L的SAEW冲洗1 min、3 min、5 min，扫描电镜（field emission scanning electron microscope，FE-SEM）观察E.f生物膜形态变化，共聚焦显微镜（confocal laser scanning microscope，CLSM）结合Live/Dead荧光染色观察E.f生物膜荧光染色情况并测定生物膜死菌比例。 结果 FE-SEM观察：50 mg/L及60 mg/L的SAEW、1% NaClO溶液随着冲洗时间的延长，生物膜中细菌减少，菌体皱缩破裂。CLSM观察：50 mg/L及60 mg/L的SAEW、1% NaClO溶液随着冲洗时间的延长，红色荧光增多，死菌比例逐步增高，三个时间点间比较差异有统计学意义（P < 0.05）；1% NaClO溶液冲洗1 min、3 min、5 min的死菌比例高于50 mg/L及60 mg/L的SAEW，差异有统计学意义（P < 0.05）。 结论 50 mg/L、60 mg/L的SAEW对根管内E. faecalis生物膜有一定的抗菌作用，但弱于1% NaClO溶液。 Abstract:Objective To study the antibacterial effect of slightly acidic electrolyzed water （SAEW） as root canal irrigation on Enterococus faecalis （E. faecalis） biofilm in root canal in vitro. Methods E. faecalis biofilms were cultured in the root canals of isolated teeth and washed with SAEW at concentrations of 50 mg/L and 60 mg/L for 1 min, 3 mins, and 5 mins, respectively, the 1% NaClO solutions was used as positive controls. The field emission scanning electron microscope （FE-SEM） was used to observe E. faecalis biofilm morphological changes, the confocal laser scanning microscope （CLSM） combined with Live/Dead fluorescence staining was used to observe the E. faecalis biofilms and analyze the proportion of dead bacteria in the biofilms. Results FE-SEM observation: 50 mg/L SAEW, 60 mg/L SAEW, 1% NaCl Osolutions with increasing washing time, E.faecalis in the biofilm decreased, the surface of the was rough, and the cells shrank. Confocal laser scanning microscope （CLSM） combined with Live/Dead fluorescence staining observation: 50 mg/L SAEW, 60 mg/L SAEW, 1% NaClO solutions with increasing washing time, the red fluorescence intensity gradually increased, and the proportion of dead bacteria gradually increased （P < 0.05）, the sterilization ratio of 1% NaClO to E. faecalis at the time point of 1 min, 3 min, and 5 min was significantly higher than 50 mg/L SAEW, 60 mg/L SAEW （P < 0.05）. Conclusion Slightly acidic electrolyzed water has a certain antibacterial effect on strains in E. faecalis biofilm, but the effect is lower than 1% NaCIO solutions. -
Key words:
- Slightly acidic electrolyzed water /
- Enterococus faecalis /
- Biofilm /
- Sodium hypochlorite /
- Antibacteria
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图 1 培养3周的E. faecalis生物膜FE-SEM及CLSM下观察
a：FE-SEM，× 10000倍；b：FE-SEM，× 50000倍；c：CLSM，× 228倍，绿色：活菌，红色：死菌，标尺为100 μm，P为髓腔侧，D为牙本质侧。
Figure 1. Observation under FE-SEM and CLSM of E. faecalis biofilm for 3 weeks
图 2 50 mg/L SAEW、60 mg/L SAEW、1% NaClO冲洗不同时间后E. faecalis生物膜FE-SEM观察
A：50 mg/L SAEW组；B：60 mg/L SAEW组；C：1% NaClO组；1：冲洗1 min；2：冲洗3 min；3：冲洗5 min；a：× 10 000倍，b：× 50000倍。
Figure 2. FE-SEM observation of E. faecalis biofilms washed with 50 mg/L SAEW，60 mg/L SAEW and 1% NaClO at different times
图 3 50 mg/L SAEW、60 mg/L SAEW、1% NaClO冲洗不同时间后CLSM荧光染色情况
（CLSM，×228倍，A：50 mg/L SAEW组、B：60 mg/L SAEW组、C：1% NaClO组；1：冲洗1 min、2：冲洗3 min、3：冲洗5 min；绿色：活菌，红色：死菌，标尺为100 μm，P为髓腔侧，D为牙本质侧）
Figure 3. E.f biofilm fluorescence staining after washed with 50 mg/L SAEW，60 mg/L SAEW and 1% NaClO at different times
表 1 不同冲洗液冲洗不同时间后的死菌比例[n = 9，（
${{\bar x}} \pm {{s}}$ ）%]Table 1. Proportion of dead bacteria washed with different washing solutions at different times [n = 9，（
${{\bar x}} \pm {{s}}$ ）%]冲洗时间 50 mg/L SAEW 60 mg/L SAEW 1% NaClO 1 min 39.33 ± 2.21*#○▲ 40.90 ± 1.84*#□ 47.41 ± 2.52*# 3 min 44.44 ± 1.64△○▲ 50.66 ± 1.61△□ 55.10 ± 3.31△ 5 min 52.39 ± 1.85○▲ 60.84 ± 2.23□ 63.57 ± 1.52 注：*表示各组1 min与3 min比较，P < 0.05；#表示各组1 min与5 min组比较，#P < 0.05；△表示各组3 min与5 min组比较，△P < 0.05；○表示50 mg/L SAEW与对照组比较，○P < 0.05；□表示60 mg/L SAEW与对照组比较，□P < 0.05；▲表示50 mg/L SAEW与60 mg/L SAEW比较，▲P < 0.05。 
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