Synergistic antibacterial study of Cu2O/CuO-tetracycline composites
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摘要:
耐药细菌快速的增长和新治疗策略的可用性越来越少，迫使人们急需研发出新型抑菌剂来解决这类难题。其中Cu2O因其抑菌效果好和细胞的毒性较小，被广泛应用于农药和防污等方面，但Cu2O自身具有离子释放过快、不稳定和易团聚等弊端限制了它的发展。tetracycline具有强的抑制活性，但细菌适应环境的能力快速增强，大大的降低了其抑菌疗效。本研究以Cu(NO3)2·3H2O为原料，水合肼为还原剂制备氧化亚铜(Cu2O/CuO)，通过与四环素(tetracycline)配位结合得到Cu2O/CuO-tetracycline复合材料。利用理论计算进一步说明Cu2O和tetracycline的配位结合方式，并探究了Cu2O/CuO-tetracycline复合材料对革兰氏阳性菌S. aureus、革兰氏阴性菌E. coli和耐药菌T-Salmonella的抑菌性能及抑菌机制。Cu2O/CuO-tetracycline复合材料具有优异的抑菌性能，进一步为公共卫生、生物医用等领域提供了广泛的科学依据。 Cu2O/CuO-tetracycline复合材料的制备流程和抑菌机制图 Abstract: The rapid growth of drug-resistant bacteria and the lack of availability of new treatment strategies make it urgent for people to develop new bacteriostatic agents to solve such problems. In this study, copper nitrate trihydrate [Cu (NO3)2·3H2O] was used as a raw material and hydrazine hydrate as a reducing agent to prepare cuprous oxide (Cu2O/CuO), then, Cu2O/CuO-tetracycline composites were obtained by combining them with tetracycline. Systematic characterization of inhibitors was conducted using transmission electron microscopy (TEM), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), Fourier- transform infrared spectroscopy (FT-IR), and ultraviolet-visible spectroscopy (UV-vis). The inhibitory properties and mechanism of Cu2O/CuO-tetracycline on the Gram-positive bacterium Staphylococcus aureus (S. aureus), Gram-negative bacterium Escherichia coli (E. coli), and drug-resistant bacterium Salmonella (T-Salmonella) were studied. The results of antibacterial properties showed that the antibacterial rate of Cu2O/CuO-tetracycline composites with antibacterial concentration of 150 µg/mL to E. coli, S. aureus, and T-Salmonella reached 99.99% at 80 min. Compared with tetracycline and Cu2O/CuO alone, the antibacterial efficiency of Cu2O/CuO-tetracycline composite increased by 2.50 and 1.38 times for E. coli, 1.58 and 1.18 times for S. aureus and 1.26 and 1.12 times for T-Salmonella, respectively. In a word, Cu2O/CuO-tetracycline composites are the most sensitive to E. coli. The antibacterial mechanism shows that the composite material can effectively destroy the bacterial cell wall, change the membrane permeability, and finally make the bacteria rupture and die. The Cu2O/CuO-tetracycline composites have excellent antibacterial properties, indicating their wide application prospects in the fields of medical devices and medical materials. -
图 1 Cu2O/CuO-tetracycline复合物的合成示意图
Figure 1. Schematic of the synthesis of Cu2O/CuO-tetracycline complexes.
图 3 (a)Cu2O/CuO和Cu2O/CuO-tetracycline的XRD谱图；Cu2O/CuO-tetracycline复合材料中Cu2 p(b)、C1 s(c)、N1 s(d)和O1 s(e)的XPS谱图；(f)tetracycline、Cu2O/CuO和Cu2O/CuO-tetracycline的FT-IR谱图；(g)光发射谱图(虚线)和Cu2O/CuO-tetracycline(紫色)和tetracycline(蓝色)的紫外可见吸收谱图(实线)
Figure 3. (a)XRD Spectra of Cu2O/CuO and Cu2O/CuO-tetracycline; XPS spectra of Cu2 p(b), C1 s(c), N1 s(d), and O1 s(e)in Cu2O/CuO-tetracycline composites; (f)FT-IR spectra of tetracycline, Cu2O/CuO, and Cu2O/CuO-tetracycline; (g)Photoemission spectra(dashed lines)and UV-vis absorption spectra(solid lines)of Cu2O/CuO-tetracycline(purple)and tetracycline(blue)
图 4 (a)tetracycline和Cu2O的优化电子结构和tetracycline的静电势(ESP)分析示意图；(b)Cu2O与tetracycline的结合能
Figure 4. (a)The optimized electronic structure of tetracycline and Cu2O and electrostatic potential (ESP) analysis diagram of tetracycline; (b)Binding Energy of Cu2O with tetracycline
图 5 在不同浓度下，tetracycline、Cu2O/CuO和Cu2O/CuO-tetracycline复合物对E. coli(a1)、S. aureus(b1)和T-Salmonella(c1)的滤纸扩散结果；E. coli(a2)、S. aureus(b2)和T-Salmonella(c2)为抑菌圈直径随浓度变化的曲线Fig. 5 At different concentrations, filter paper diffusion results of tetracycline, Cu2O/CuO and Cu2O/CuO-tetracycline composites on E. coli (a1), S. aureus (b1) and T-Salmonella (c1); E. coli(a2)、S. aureus(b2)和T-Salmonella(c2)are the curves of the diameter of inhibition zone changing with concentration.
图 6 浓度为150 µg/mL的Cu2O/CuO-tetracycline复合材料对E. coli(a)、S. aureus(b)和T-Salmonella(c)的菌落计数结果；(d)Cu2O/CuO-tetracycline在不同时间对三种测试菌的抑菌率；(e)Cu2O/CuO-tetracycline在不同时间对三种测试菌的菌落数；(f)浓度为150 µg/mL的Cu2O/CuO-tetracycline复合材料与三种测试菌混合5和40 min后的zeta电位值
Figure 6. Colony counting results of E. coli(a), S. aureus(b)and T-Salmonella(c)by Cu2O/CuO-tetracycline composites with concentration of 150 µg/mL;(d)Antibacterial rate of Cu2O/CuO-tetracycline compound to three experimental bacteria in different time; (e)Colony number of Cu2O/CuO-tetracycline composites to three kinds of tested bacteria in different time; (f)zeta potential value of Cu2O/CuO-tetracycline composites with concentration of 150 µg/mL mixed with three experimental bacteria for 5 and 40 min.
图 7 (A)ICP-OES测得铜阳离子的累积释放;(B)(a1~3)分别为空白对照组纯菌E. coli、S. aureus和T-Salmonella的代表性荧光图像, (b1~3)分别为Cu2O/CuO-tetracycline复合材料与E. coli、S. aureus和T-Salmonella接触2 h后的代表性荧光图像
Figure 7. (A) Cumulative release of copper cations measured by ICP-OES; (B) (a1~3) are representative fluorescence images of pure bacteria E. coli, S. aureus and T-Salmonella in the blank control group respectively，(b1~3) are the representative fluorescence images of Cu2O/CuO-tetracycline composite after contacting with E. coli, S. aureus and T-Salmonella for 2 h, respectively.
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