二氧化钛纳米粒子联合LED光照射对口腔变异链球菌的抑制作用及其机制

doi:10.13481/j.1671-587X.20220412

吉林大学学报(医学版) ›› 2022, Vol. 48 ›› Issue (4): 929-937.doi: 10.13481/j.1671-587X.20220412

二氧化钛纳米粒子联合LED光照射对口腔变异链球菌的抑制作用及其机制

闫琳琳^1,²,朱琳³,赵远航^1,²,宋嘉卓^1,²,邹馨颖^1,²,刘新^1,²,赵红^1,²,张志民¹(),张红¹()

^1.吉林大学口腔医院牙体牙髓科，吉林长春 130021
^2.吉林省口腔生物医学国际联合研究中心，吉林长春 130021
^3.吉林大学超分子结构与材料国家重点实验室，吉林长春 130012

收稿日期:2021-11-24 出版日期:2022-07-28 发布日期:2022-07-26
通讯作者: 张志民,张红 E-mail:zhangzhim@jlu.edu.cn;zhanghong1983@jlu.edu.cn
作者简介:闫琳琳（1987－），女，陕西省汉中市人，在读硕士研究生，主要从事口腔抗菌材料方面的研究。
基金资助:
吉林省发改委2020年省预算内基本建设资金项目(2020C029-2);吉林省财政厅科技项目(JCSZ2020304-2)

Inhibitory effect of Streptococcus mutans by titanium dioxide nanoparticles combined with LED light irradiation and its mechanism

Linlin YAN^1,²,Lin ZHU³,Yuanhang ZHAO^1,²,Jiazhuo SONG^1,²,Xinying ZOU^1,²,Xin LIU^1,²,Hong ZHAO^1,²,Zhimin ZHANG¹(),Hong ZHANG¹()

^1.Department of Endodontics，Stomatology Hospital，Jilin University，Changchun 130021，China
^2.Jilin Provincial International Cooperation Base of Oral Biomedicine，Changchun 130021，China
^3.State Key Laboratory of Supramolecular Structure and Materials，Jilin University，Changchun 130021，China

Received:2021-11-24 Online:2022-07-28 Published:2022-07-26
Contact: Zhimin ZHANG,Hong ZHANG E-mail:zhangzhim@jlu.edu.cn;zhanghong1983@jlu.edu.cn

摘要/Abstract

摘要： 目的

探讨对二氧化钛纳米粒子（TiO₂NPs）进行锌（Zn）和氮（N）共掺杂时最适掺杂浓度及Zn和N共掺杂TiO₂NPs（Zn-N-TiO₂NPs）对口腔变异链球菌的抑制作用，阐明其作用机制。

方法

采用钛酸丁酯、硝酸锌和氨水分别作为钛（Ti）源、Zn源和N源，利用溶胶-水热法合成TiO₂NPs和不同掺杂浓度的Zn-N-TiO₂NPs，通过X射线衍射（XRD）、拉曼光谱、扫描电子显微镜（SEM）和紫外-可见光吸收光谱分析（UV-vis）对纳米粒子进行表征，采用Cure Rite辐射仪对牙科发光二极管（LED）光固化灯进行表征。变异链球菌分为空白对照组、单独LED光照组、TiO₂NPs组、1%Zn-3%N-TiO₂NPs（Zn1）组、3%Zn-3%N-TiO₂NPs（Zn3）组、5%Zn-3%N-TiO₂NPs（Zn5）组和7%Zn-3%N-TiO₂NPs（Zn7）组。将各组纳米粒子与变异链球菌菌液混合（终浓度为2 g·L^－1）摇匀。除空白对照组，其他各组采用牙科LED光分别照射1、3和5 min，空白对照组仅用培养基处理，采用平板菌落计数法记录各组平板菌落数。选取抗菌效果最佳组（Zn3组），利用1，3-二苯基异苯并呋喃（DPBF）探针检测活性氧（ROS）释放量，分为0 min组、1 min组、3 min组和5 min组，每组取2 g· L^－1的Zn3-DPBF溶液100 μL分别采用牙科LED光照0、1、3和5 min，观察各组溶液在波长410 nm处吸收峰值，即代表ROS释放量。变异链球菌分为对照组、Ｎ-乙酰半胱氨酸（NAC）组、Zn3组和NAC+Zn3组，LED光照射5 min后避光培养24 h，采用平板菌落计数法计数各组平板菌落数。

结果

成功合成Zn-N-TiO₂NPs，晶体结构为锐钛矿型，TiO₂NPs组、Zn1组、Zn3组、Zn5组和Zn7组纳米粒子平均晶粒尺寸分别为15.6、11.3、9.8、9.4和7.3 nm；SEM观察纳米粒子为分布均匀的球形颗粒；UV-vis显示Zn3在400~500 nm波长范围内吸光度（A）值明显升高。与空白对照组比较，LED光照射5 min时Zn3组菌落数最少（P<0.05）。与0 min组比较，1 min组、3 min组和5 min组Zn3-DPBF溶液在波长410 nm处吸收峰值依次下降，表明各组纳米粒子溶液ROS释放量随光照时间延长而逐渐增加。与Zn3组比较，NAC+Zn3组菌落数明显增加（P<0.05）。

结论

Zn和N掺杂浓度均为3%的TiO₂NPs（3%Zn-3%N-TiO₂NPs）联合LED光照射通过光催化作用能有效抑制口腔变异链球菌的生长。

关键词: 二氧化钛, 掺杂, 变异链球菌, 抗菌性能

Abstract: Objective

To investigate the optimum doping concentrations of zinc（Zn） and nitrogen（N） in co-doping of titanium dioxide nanoparticles（TiO₂NPs） and the inhibitory effect of co-doping of Zn， N and TiO₂NPs（Zn-N-TiO₂NPs） on the Streptococcus mutans， and to clarify its mechanism.

Methods

The TiO₂NPs and Zn-N-TiO₂NPs with different doping concentrations were synthesized by sol-hydrothermal method using butyl titanate， zinc nitrate and ammonia as titanium source，Zn source and N source，respectively.X-ray diffraction（XRD），Raman spectra，scanning electron microscope（SEM），UV-visible absorption spectroscope（UV-vis） were used to characterize the nanoparticles in various groups.Cure Rite radiometer was used to characterize the dental light emitting diode curing lamp.The Streptococcus mutans were divided into blank control group，LED light irradiation group，TiO₂NPs group，1%Zn-3%N-TiO₂NPs group（Zn1 group），3%Zn-3%N-TiO₂NPs group（Zn3 group），5%Zn-3%N-TiO₂NPs group（Zn5 group），and 7%Zn-3%N-TiO₂NPs group（Zn7 group）.The nanoparticles in each group were mixed with Streptococcus mutans bacteria solution（the final concentration of nanoparticles was 2 g·L^－1） and were shaken well.The nanoparticles groups and the LED light irradiation group were exposed to dental LED light for 1， 3，and 5 min，the blank control group was treated with culture medium only，and the number of colonies in each group was recorded by plate colony counting method.The group with the best antibacterial effect（Zn3 group） was selected.The reactive oxygen species（ROS） release amount was detected using DPBF probe，and the experiment was divided into 0 min group， 1 min group， 3 min group and 5 min group.A total of 100 μL Zn3-DPBF solution at the concentration of 2 g·L^－1 in each group was selected for dental LED light irradiation for 0， 1， 3 and 5 min， respectively， and the changes in absorption peaks at 410 nm were observed，indicating the ROS release amount.In the experiment exploring the influence of N-acetyl cystethione（NAC） in the inhibitory effect of Zn3 combined with LED light irradiation on Streptococcus mutans，Streptococcus mutans were divided into control group， 5 mmol·L^－1NAC group，Zn3 group and NAC +Zn3 group.After LED light irradiation for 5 min and culture avoiding light for 24 h，plate colony counting method was used to record the number of colonies in each group.

Results

Zn-N-TiO₂NPs was successfully synthesized and its crystal structure was anatase.The average sizes of nanoparticles in TiO₂NPs group，Zn3 group，Zn5 group and Zn7 group were 15.6， 11.3， 9.8， 9.4 and 7.3 nm， respectively.The SEM results showed that the nanoparticles were spherical particles with uniform distribution.The Uv-vis results showed that the absorbance（A） value of Zn3 was increased significantly in the wavelength range of 400－500 nm.Compared with blank control group， the colony count in Zn3 group was the least when LED irradiation was 5 min（P<0.05）.Compared with 0 min group，the absorption peaks of Zn3-DPBF solution at 410 nm in 1 min，3 min and 5 min groups were decreased successively， indicating the ROS relaease amount of nanoparticle solution in various were increased with the prolongation of illumination time.Compared with Zn3 group，the number of colonies in NAC+Zn3 group was significantly increased（P<0.05）.

Conclusion

3%Zn-3%N-TiO₂NPs combined with LED light irradiation can effectively inhibit the growth of the Streptococcus mutans through photocatalysis.

Key words: Titanium dioxide, Doping, Streptococcus mutans, Antibacterial property

中图分类号:

R781.1

闫琳琳,朱琳,赵远航,宋嘉卓,邹馨颖,刘新,赵红,张志民,张红. 二氧化钛纳米粒子联合LED光照射对口腔变异链球菌的抑制作用及其机制[J]. 吉林大学学报(医学版), 2022, 48(4): 929-937.

Linlin YAN,Lin ZHU,Yuanhang ZHAO,Jiazhuo SONG,Xinying ZOU,Xin LIU,Hong ZHAO,Zhimin ZHANG,Hong ZHANG. Inhibitory effect of Streptococcus mutans by titanium dioxide nanoparticles combined with LED light irradiation and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2022, 48(4): 929-937.

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参考文献 29

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Group	Number of colonies	Group	Number of colonies
Blank control	530.00±17.80^*	Zn3
LED illumination		LED 1 min	18.67±3.30^△＃
LED 1 min	525.33±16.68^*	LED 3 min	7.67±2.05^△＃
LED 3 min	483.67±7.72^*△	LED 5 min	3.00±1.63^△＃
LED 5 min	444.00±13.14^*△	Zn5
TiO₂NPs		LED 1 min	97.67±4.92^*△＃
LED 1 min	248.33±15.52^*△＃	LED 3 min	66.00±3.74^*△＃
LED 3 min	193.67±13.47^*△＃	LED 5 min	43.33±4.64^*△＃
LED 5 min	157.67±11.90^*△＃	Zn7
Zn1		LED 1 min	131.00±3.56^*△＃
LED 1 min	44.67±4.78^*△＃	LED 3 min	116.33±3.09^*△＃
LED 3 min	38.00±3.74^*△＃	LED 5 min	106.33±4.99^*△＃
LED 5 min	25.33±7.76^*△＃

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二氧化钛纳米粒子联合LED光照射对口腔变异链球菌的抑制作用及其机制

Inhibitory effect of Streptococcus mutans by titanium dioxide nanoparticles combined with LED light irradiation and its mechanism

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