具有多酶活性CuO/ZnO复合纳米球的制备及其抗菌性能研究

doi:10.6023/A25050176

摘要/Abstract

细菌感染极大威胁着人类的健康, 由于抗生素的长期滥用而产生的“超级细菌”甚至让人类处于近乎“无药可用”的境地. 基于纳米酶的催化疗法为耐药菌的感染治疗提供了一种新策略. 然而, 单一的催化活性和有限的抗菌效率极大地限制了其进一步发展. 以聚丙烯酸钠纳米球(PAAS NSs)为模板, 成功制备了具有多酶活性的CuO/ZnO复合纳米球(CuO/ZnO NSs). 通过3,3',5,5'-四甲基联苯胺(TMB)和邻苯二胺(OPDA)两种探针证实了CuO/ZnO NSs具有类氧化物酶(OXD-like)活性和类过氧化物酶(POD-like)活性, 可在弱酸性条件下催化O₂和H₂O₂生成大量的活性氧(ROS). 平板计数实验结果表明, 在H₂O₂存在时, 50 μg/mL的CuO/ZnO NSs对大肠杆菌(E. coli)和耐甲氧西林金黄色葡萄球菌(MRSA)均有100%的抑制率. 对CuO/ZnO NSs的抗菌机制研究发现破坏细菌膜和诱导内部蛋白泄露是其主要抗菌机理. 此外, CCK-8实验证明CuO/ZnO NSs具有良好的生物安全性. 因此, 本研究为金属氧化物类纳米酶在细菌感染治疗领域的应用提供了新的研究思路.

关键词: 复合纳米球, 多酶活性, 细菌感染, 活性氧, 抗菌机理

Bacterial infections pose a significant threat to human health. Clinically, the treatment of bacterial infections often relies on antibiotics. However, the emergence of “multi-drug resistant bacteria” due to antibiotic misuse has been a persistent challenge for the medical community, even leading humanity to a near “antibiotic-free” predicament. Metal-based nanozymes, with their high antibacterial activity and low resistance, have emerged as effective alternative strategies for treating bacterial infection wounds. However, single catalytic activity and limited antibacterial effects restrict its further development. In this study, a CuO/ZnO composite nanosphere was successfully prepared and used as nanozyme for the eradication of antibiotic-resistant bacteria. The CuO/ZnO composite nanosphere was characterized using transmission electron microscopy, scanning electron microscopy, X-ray diffraction, X-ray photoelectron spectroscopy, and nitrogen adsorption instruments, confirming its successful synthesis. The catalytic activity of the material was evaluated using two reactive oxygen probes, 3,3',5,5'-tetramethylbenzidine (TMB) and o-phenylenediamine (OPDA), revealing that the CuO/ZnO composite nanospheres possess both oxidase-like (OXD-like) and peroxidase-like (POD-like) activities. The catalytic activity of the composite nanospheres is significantly higher than that of individual CuO and ZnO nanospheres, validating the rationality of our synthesis strategy. The antibacterial performance of CuO/ZnO composite nanospheres at different concentrations was tested using the viable cell count method. The results indicated that the CuO/ZnO composite nanospheres at a concentration of 50 μg/mL, in the presence of H₂O₂, could generate reactive oxygen species through their multi-enzyme activity, effectively killing Escherichia coli (E. coli) and methicillin-resistant Staphylococcus aureus (MRSA), achieving a 100% inhibition rate. Subsequent studies on the antibacterial mechanism revealed that the high-efficiency bactericidal effect of CuO/ZnO composite nanospheres is primarily attributed to their disruption of bacterial cell membranes and leakage of intracellular proteins. Additionally, CCK-8 assays confirmed the good biocompatibility of CuO/ZnO composite nanosphere, with cell viability remaining above 80% even at a concentration of 100 μg/mL. In summary, this study provides new research perspectives for the application of metal-based nanozymes in the treatment of drug-resistant bacterial infections.

Key words: composite nanosphere, multi-enzyme activity, bacterial infection, reactive oxygen species, antibacterial mechanism

引用此文

施寒竹, 高雅宁, 陈昭玉, 戴陈伟, 周秀红, 张颖颖. 具有多酶活性CuO/ZnO复合纳米球的制备及其抗菌性能研究[J]. 化学学报, 2025, 83(11): 1363-1371.

Shi Hanzhu, Gao Yaning, Chen Zhaoyu, Dai Chenwei, Zhou Xiuhong, Zhang Yingying. Preparation of CuO/ZnO Composite Nanospheres with Multi-enzyme Activity and Study of Their Antibacterial Properties[J]. Acta Chimica Sinica, 2025, 83(11): 1363-1371.

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图/表 7

图式1. CuO/ZnO纳米球的合成及应用示意图

Scheme 1. Schematic diagram of the synthesis and application of CuO/ZnO nanospheres

图1. CuO/ZnO纳米球的(A)透射电镜图像(TEM)、(B)扫描电镜图像(SEM)、(C)高分辨率SEM图像及(D)高分辨TEM图像

Figure 1. (A) Transmission electron microscopy (TEM) image, (B) scanning electron microscopy (SEM) image, (C) high-resolution SEM image and (D) high-resolution TEM image of CuO/ZnO nanospheres

图2. (A) CuO/ZnO纳米球的傅里叶红外光谱图、(B) CuO/ZnO纳米球的X射线光电子能谱图(XPS)和(C, D) CuO/ZnO纳米球的高分辨XPS能谱图[Cu 2p (C), Zn 2p (D)]

Figure 2. (A) Fourier-transform infrared spectroscopy (FTIR) spectrum of CuO/ZnO nanospheres, (B) X-ray photoelectron spectroscopy (XPS) spectrum of CuO/ZnO nanospheres and (C, D) High-resolution XPS spectra of CuO/ZnO nanospheres [Cu 2p (C), Zn 2p (D)]

图3. (A) CuO/ZnO纳米球的类氧化酶活性(OXD-like)和类过氧化物酶(POD-like)活性示意图、使用TMB探针得到的不同组合(B)、不同浓度(C)及不同H2O2浓度(D)下的CuO/ZnO纳米球的类酶催化活性以及使用OPDA探针得到的不同组合(E)、不同浓度(F)及不同H2O2浓度(G)下的CuO/ZnO纳米球的类酶催化活性

Figure 3. (A) Schematic representation of the OXD-like and POD-like enzymatic activities of CuO/ZnO nanospheres, enzyme-like catalytic activity of CuO/ZnO nanospheres obtained under different combinations (B), concentrations (C) and H2O2 concentrations (D) using TMB probe, and enzyme-like catalytic activity of CuO/ZnO nanospheres obtained under different combinations (E), concentrations (F) and H2O2 concentrations (G) using OPDA prob

图4. 有无H2O2存在时不同浓度的CuO/ZnO纳米球处理后(A) E. coli和(B) MRSA在平板上的菌落数以及(C) E. coli和(D) MRSA的细菌存活率统计图

Figure 4. (A) Number of colonies of E. coli (A) and MRSA (B) on plates and statistical graph of the lower bacterial survival rate of (C) E. coli and (D) MRSA after CuO/ZnO nanosphere treatment after treatment with CuO/ZnO nanospheres at different concentrations, with and without the presence of H2O

图5. 有无H2O2存在时不同浓度的CuO/ZnO纳米球处理后(A) E. coli和(B) MRSA的扫描电镜(SEM)图像(箭头为细菌破损处)及(C) E. coli和(D) MRSA的蛋白泄露统计图

Figure 5. Scanning electron microscopy (SEM) images of E. coli (A) and MRSA (B) (arrows indicate bacterial damage sites) and protein leakage statistics of E. coli (C) and MRSA (D) after treatment with CuO/ZnO nanoparticles of different concentrations, with or without the presence of H2O2

图6. 不同浓度的CuO/ZnO纳米球与(A)人脐静脉内皮细胞(HUVECs)及(B)小鼠成纤维细胞(L929)培养24 h后的细胞存活率统计图

Figure 6. Statistical results of cell survival after 24 h culture of different concentrations of CuO/ZnO nanospheres with (A) human umbilical vein endothelial cells (HUVECs) and (B) mouse fibroblasts (L929)

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