TiO2纳米管负载黄芩苷-壳聚糖复合涂层的生物学评价及抗菌性能研究

doi:10.13591/j.cnki.kqyx.2025.12.004

摘要/Abstract

摘要：

目的研究TiO₂纳米管负载黄芩苷-壳聚糖复合涂层的生物学效果及抗菌性能。方法 ①在钛片表面制备TiO₂纳米管,将其分为4组:阳极氧化组(TiO₂)、阳极氧化载黄芩苷组(TiO₂+BC)、阳极氧化载壳聚糖组(TiO₂+CS)和阳极氧化载黄芩苷-壳聚糖复合涂层组(TiO₂+BC+CS)。通过扫描电镜、表面接触角以及高效液相色谱实验来检测各组理化性能。②将大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)与各组钛片共培养,通过CCK-8黏附增殖实验、活/死细胞染色、ALP活性检测、茜素红S染色及细胞矿化定量检测评价黄芩苷-壳聚糖复合涂层对BMSCs黏附、增殖和成骨向分化的影响。③将各组钛片与金黄色葡萄球菌(S.aureus)和大肠杆菌(E.coli)共培养,通过抑菌环实验、平板涂布法、浊度法来测试该涂层的抗菌性能。结果 ①扫描电镜下观测到壳聚糖可覆盖TiO₂纳米管并在其表面形成一层膜状结构,接触角测试结果显示TiO₂+BC+CS组接触角大于TiO₂组(P<0.01),亲水性降低,但与TiO₂+BC组接触角无差异(P>0.05)。高效液相色谱法测得TiO₂+BC+CS可延缓黄芩苷的释放,达到缓释效果。②细胞黏附实验表明4 h时TiO₂+BC+CS组细胞数目多于TiO₂组(P<0.05);CCK-8增殖检测表明7 d时TiO₂+BC+CS组细胞增殖速度快于TiO₂组和TiO₂+BC组(P<0.05);活/死细胞染色结果显示各组钛片表面均附着大量绿色荧光活细胞,细胞形态类似。4 d时TiO₂+BC组和TiO₂+BC+CS组ALP活性较高,7 d时TiO₂+BC+CS组ALP活性最高(P<0.05)。茜素红S染色结果显示TiO₂+BC+CS组表面红染颗粒最多,颜色更深,在14、21 d时,TiO₂+BC+CS组的细胞矿化水平均最高(P<0.05)。③TiO₂+BC+CS组周围出现明显抑菌环,TiO₂+CS组和TiO₂+BC+CS组菌液浊度低于TiO₂组和TiO₂+BC组(P<0.05)。结论 TiO₂纳米管负载黄芩苷-壳聚糖复合涂层有利于BMSCs的早期黏附、增殖和成骨分化,并具有良好的抗菌性能。

关键词: 黄芩苷, 壳聚糖, 生物学评价, 抗菌性能

Abstract:

Objective To investigate the biological effects and antibacterial properties of TiO₂ nanotubes loaded baicalin-chitosan composite coating. Methods ①TiO₂ nanotubes were fabricated on the surface of titanium sheet and divided into four groups:TiO₂, TiO₂+BC, TiO₂+CS, TiO₂+BC+CS. The physicochemical properties of each group were detected by scanning electron microscope, surface contact angle and high performance liquid chromatography. ②BMSCs were co-cultured with titanium tablets of each group, and the effects of baicalin-chitosan composite coating on adhesion, proliferation and osteogenic differentiation of BMSCs were evaluated by CCK-8 adhesion proliferation test, live/dead cell staining, ALP activity detection, alizarin red S staining and cell mineralization quantitative detection. ③The titanium sheets of each group were co-cultured with S. aureus and E. coli, and the antibacterial properties of the coating were tested by zone of inhibition test, spread plate method and turbidimetric method. Results ①Under scanning electron microscope, it was observed that chitosan could cover the TiO₂ nanotubes and form a membrane-like structure on the surface. The contact angle test results showed that the contact angle of TiO₂+BC+CS group was larger than that of TiO₂ group (P<0.01), and the hydrophilicity was worse, but there was no difference with TiO₂+BC group (P>0.05). High performance liquid chromatography (HPLC) showed that TiO₂+BC+CS group could delay the release of baicalin and achieve a slow release effect. ②BMSCs were co-cultured with titanium tablets of each group. The cell adhesion test showed that the number of cells in TiO₂+BC+CS group was higher than that in TiO₂ group at 4 h (P<0.05). CCK-8 proliferation detection showed that the cell proliferation rate of TiO₂+BC+CS group was faster than that of TiO₂+BC group and TiO₂+BC group at 7 days (P<0.05). The staining results of live/dead cells showed that there were a large number of green fluorescent live cells that attached to the surface of each group, and the cell morphology was similar. The ALP activity of TiO₂+BC and TiO₂+BC+CS groups was higher at 4 d, and the ALP activity of TiO₂+BC+CS group was the best at 7 d (P<0.05). The results of alizarin red S staining showed that TiO₂+BC+CS group had the most red particles and the color was darker. At 14 and 21 days, the cell mineralization level of TiO₂+BC+CS group was the highest (P<0.05). ③TiO₂+BC+CS group had obvious inhibition zone around it, and the turbidity of bacterial solution in TiO₂+CS group and TiO₂+BC+CS group was lower than that in TiO₂ group and TiO₂+BC group (P<0.05). Conclusion The baicalin-chitosan composite coating is beneficial to the early adhesion, proliferation and osteogenic differentiation of BMSCs and has good antibacterial properties.

Key words: baicalin, chitosan, biological evaluation, antibacterial properties

中图分类号:

R780.2

张婷婷, 李潇, 冯大军, 马燕, 谈珺, 刘莉. TiO₂纳米管负载黄芩苷-壳聚糖复合涂层的生物学评价及抗菌性能研究[J]. 口腔医学, 2025, 45(12): 901-908.

ZHANG Tingting, LI Xiao, FENG Dajun, MA Yan, TAN Jun, LIU Li. Biological evaluation and study on antibacterial properties of TiO₂ nanotubes loaded baicalin-chitosan composite coating[J]. Stomatology, 2025, 45(12): 901-908.

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组别	2 h	8 h	12 h	24 h	72 h	120 h	168 h
TiO₂+BC组	17.170±2.129	6.790±1.187	0.561±0.207	0.500±0.278	0.164±0.132	0	0
TiO₂+BC+CS组	11.288±2.689	5.597±0.487	2.511±0.050	1.581±0.271	0.919±0.182	0.305±0.097	0.222±0.101
P	0.041	0.183	0.001	0.009	0.004

TiO₂纳米管负载黄芩苷-壳聚糖复合涂层的生物学评价及抗菌性能研究

Biological evaluation and study on antibacterial properties of TiO₂ nanotubes loaded baicalin-chitosan composite coating

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