Ag/TiO_2纳米材料对烟曲霉的抑制作用及机制
|
摘要
目的合成Ag/TiO_2纳米材料,对其进行表征测定,并探讨其对烟曲霉的抑制作用及具体机制。方法采用光催化还原法制备Ag/TiO_2纳米材料,紫外可见分析和扫描电镜对其进行表征测定;微量液基稀释法检测对烟曲霉的最低抑菌浓度(MIC),以及生物量的抑制作用;ELISA试剂盒检测对真菌谷胱甘肽还原酶、总谷胱甘肽、线粒体膜电位的影响,荧光显微镜检测活性氧的产生。结果成功制备Ag/TiO_2纳米材料,分布均匀;对烟曲霉的MIC值为0.5μg/mL,能完全抑制烟曲霉生物量,与单独纳米银相比,具有更好的抗菌活性。机制研究发现其主要通过降低烟曲霉体内谷胱甘肽及其还原酶的含量,诱导过量活性氧的产生,最终导致线粒体膜电位降低,使真菌细胞发生凋亡。结论 Ag/TiO2纳米材料可有效阻断烟曲霉等真菌在空气中的传播,具有广泛的应用前景。
Objective To synthesize and characterize Ag/TiO2 nanomaterials,and explore their antifungal action and specific mechanism against Aspergillus fumigatus Fresenius.Methods Ag/TiO2 nanomaterials were prepared with photocatalytic reduction method;UV-vis analysis and scanning electron microscopy were used to characterize them.The minimum inhibitory concentration(MIC)of Ag/TiO2 nanomaterials and the inhibitory effect on biomass were detected with microdilution method.The ELISA kits were used to detect their effects on fungal glutathione reductase,total glutathione and mitochondrial membrane potential.The production of reactive oxygen was detected by using fluorescence microscopy.Results The Ag/TiO2 nanomaterials were successfully prepared,which distributed evenly.The MIC value of Ag/TiO2 nanomaterials against Aspergillus fumigatus Fresenius was 0.5μg/mL.The materials completely inhibited the biomass,and showed better antibacterial activity compared with the single nano silver.Mechanism study found that it induced the production of excess reactive oxygen mainly by reducing the contents of glutathione and its reductase in Aspergillus fumigatus Fresenius,which eventually led to a decrease in mitochondrial membrane potential and apoptosis of fungal cells.Conclusion Ag/TiO2 nanomaterials can effectively block the spread of Aspergillus fumigatus Fresenius and other fungi in the air,which has a wide application prospect.
Objective To synthesize and characterize Ag/TiO2 nanomaterials,and explore their antifungal action and specific mechanism against Aspergillus fumigatus Fresenius.Methods Ag/TiO2 nanomaterials were prepared with photocatalytic reduction method;UV-vis analysis and scanning electron microscopy were used to characterize them.The minimum inhibitory concentration(MIC)of Ag/TiO2 nanomaterials and the inhibitory effect on biomass were detected with microdilution method.The ELISA kits were used to detect their effects on fungal glutathione reductase,total glutathione and mitochondrial membrane potential.The production of reactive oxygen was detected by using fluorescence microscopy.Results The Ag/TiO2 nanomaterials were successfully prepared,which distributed evenly.The MIC value of Ag/TiO2 nanomaterials against Aspergillus fumigatus Fresenius was 0.5μg/mL.The materials completely inhibited the biomass,and showed better antibacterial activity compared with the single nano silver.Mechanism study found that it induced the production of excess reactive oxygen mainly by reducing the contents of glutathione and its reductase in Aspergillus fumigatus Fresenius,which eventually led to a decrease in mitochondrial membrane potential and apoptosis of fungal cells.Conclusion Ag/TiO2 nanomaterials can effectively block the spread of Aspergillus fumigatus Fresenius and other fungi in the air,which has a wide application prospect.
引文
[1]Durán N,Marcato PD,De Souza GIH,et al.Antibacterial effect of silver nanoparticles produced by fungal process on textile fabrics and their effluent treatment[J].J Biomed Nanotechnol,2007,3(2):203-208.
[2]Onnis V,De LA,Cocco MT,et al.2-Acylhydrazino-5-arylpyrrole derivatives:Synthesis and antifungal activity evaluation[J].Eur JMed Chem,2009,44(3):1288-1295.
[3]XU Weiping.Research on composite nano-antibacterial materials based on silver[D].Peking:University of Science and Technology of China,2011.(in Chinese)徐维平.基于银的复合纳米抗菌材料的研究[D].北京:中国科学技术大学,2011.
[4]Ul-Islam M,Shehzad A,Khan S,et al.Antimicrobial and biocompatible properties of nanomaterials[J].J Nanosci&Nanotechnol,2014,14(1):780-791.
[5]Rai M,Birla S,Ingle AP,et al.Nanosilver:An inorganic nanoparticle with myriad potential applications[J].Nanotechnol Rev,2014,3(3):281-309.
[6]Muhsin TM,Hachim AK.Mycosynthesis and characterization of silver nanoparticles and their activity against some human pathogenic bacteria[J].World J Microbiol&Biotechnol,2014,30(7):2081-2090.
[7]Paredes D,Ortiz C,Torres R.Synthesis,characterization,and evaluation of antibacterial effect of Ag nanoparticles against Escherichia coli O157:H7and methicillin-resistant Staphylococcus aureus(MRSA)[J].Int J Nanomed,2014,9(Issue 1):1717-1729.
[8]Kim KJ,Sung WS,Moon SK,et al.Antifungal effect of silver nanoparticles on dermatophytes[J].J Microbiol&Biotechnol,2008,18(8):1482-1484.
[9]Zhang Y,Peng H,Huang W,et al.Facile preparation and characterization of highly antimicrobial colloid Ag or Au nanoparticles[J].J Colloid&Interface Sci,2008,325(2):371-376.
[10]Chen SG,Chen SJ,Jiang S,et al.Study of zwitterionic sulfopropylbetaine containing reactive siloxanes for application in antibacterial materials[J].Colloids&Surfaces B:Biointerfaces,2011,85(2):323-329.
[11]Rao KJ,Paria S.Use of sulfur nanoparticles as a green pesticide on Fusarium solani and Venturia inaequalis phytopathogens[J].RSC Adv,2013,3(26):10471-10478.
[12]Ott M,Gogvadze V,Orrenius S,et al.Mitochondria,oxidative stress and cell death[J].Apoptosis,2007,12(5):913-922.
[13]SHI Yuping.Biological function analysis of glutathione reductase gene in Acidithiobacillus caldus MTH-04[D].Ji′nan:Shandong University,2015.(in Chinese)施玉萍.嗜酸性喜温硫杆菌谷胱甘肽还原酶基因的生物学功能研究[D].济南:山东大学,2015.
[14]YAN Huifang,MAO Peisheng,XIA Fangshan.Advances in the study of glutathione,aplant antioxidant[J].Grassland J,2013,21(3):428-434(in Chinese).闫慧芳,毛培胜,夏方山.植物抗氧化剂谷胱甘肽研究进展[J].草地学报,2013,21(3):428-434.
[15]Xia Z,Lundgren B,Bergstrand A,et al.Changes in the generation of reactive oxygen species and in mitochondrial membrane potential during apoptosis induced by the antidepressants imipramine,clomipramine,and citalopram and the effects on these changes by Bcl-2and Bcl-XL[J].Biochem Pharmacol,1999,57(10):1199-1208.
[16]Bink A,Vandenbosch D,Coenye T,et al.Superoxide dismutases are involved in Candida albicans biofilm persistence against miconazole[J].Antimicrob Agents Chemother,2011,55(9):4033-4037.
[17]Delattin N,Cammue BP,Thevissen K.Reactive oxygen species-inducing antifungal agents and their activity against fungal biofilms[J].Fut Med Chem,2014,6(1):77-90.
[2]Onnis V,De LA,Cocco MT,et al.2-Acylhydrazino-5-arylpyrrole derivatives:Synthesis and antifungal activity evaluation[J].Eur JMed Chem,2009,44(3):1288-1295.
[3]XU Weiping.Research on composite nano-antibacterial materials based on silver[D].Peking:University of Science and Technology of China,2011.(in Chinese)徐维平.基于银的复合纳米抗菌材料的研究[D].北京:中国科学技术大学,2011.
[4]Ul-Islam M,Shehzad A,Khan S,et al.Antimicrobial and biocompatible properties of nanomaterials[J].J Nanosci&Nanotechnol,2014,14(1):780-791.
[5]Rai M,Birla S,Ingle AP,et al.Nanosilver:An inorganic nanoparticle with myriad potential applications[J].Nanotechnol Rev,2014,3(3):281-309.
[6]Muhsin TM,Hachim AK.Mycosynthesis and characterization of silver nanoparticles and their activity against some human pathogenic bacteria[J].World J Microbiol&Biotechnol,2014,30(7):2081-2090.
[7]Paredes D,Ortiz C,Torres R.Synthesis,characterization,and evaluation of antibacterial effect of Ag nanoparticles against Escherichia coli O157:H7and methicillin-resistant Staphylococcus aureus(MRSA)[J].Int J Nanomed,2014,9(Issue 1):1717-1729.
[8]Kim KJ,Sung WS,Moon SK,et al.Antifungal effect of silver nanoparticles on dermatophytes[J].J Microbiol&Biotechnol,2008,18(8):1482-1484.
[9]Zhang Y,Peng H,Huang W,et al.Facile preparation and characterization of highly antimicrobial colloid Ag or Au nanoparticles[J].J Colloid&Interface Sci,2008,325(2):371-376.
[10]Chen SG,Chen SJ,Jiang S,et al.Study of zwitterionic sulfopropylbetaine containing reactive siloxanes for application in antibacterial materials[J].Colloids&Surfaces B:Biointerfaces,2011,85(2):323-329.
[11]Rao KJ,Paria S.Use of sulfur nanoparticles as a green pesticide on Fusarium solani and Venturia inaequalis phytopathogens[J].RSC Adv,2013,3(26):10471-10478.
[12]Ott M,Gogvadze V,Orrenius S,et al.Mitochondria,oxidative stress and cell death[J].Apoptosis,2007,12(5):913-922.
[13]SHI Yuping.Biological function analysis of glutathione reductase gene in Acidithiobacillus caldus MTH-04[D].Ji′nan:Shandong University,2015.(in Chinese)施玉萍.嗜酸性喜温硫杆菌谷胱甘肽还原酶基因的生物学功能研究[D].济南:山东大学,2015.
[14]YAN Huifang,MAO Peisheng,XIA Fangshan.Advances in the study of glutathione,aplant antioxidant[J].Grassland J,2013,21(3):428-434(in Chinese).闫慧芳,毛培胜,夏方山.植物抗氧化剂谷胱甘肽研究进展[J].草地学报,2013,21(3):428-434.
[15]Xia Z,Lundgren B,Bergstrand A,et al.Changes in the generation of reactive oxygen species and in mitochondrial membrane potential during apoptosis induced by the antidepressants imipramine,clomipramine,and citalopram and the effects on these changes by Bcl-2and Bcl-XL[J].Biochem Pharmacol,1999,57(10):1199-1208.
[16]Bink A,Vandenbosch D,Coenye T,et al.Superoxide dismutases are involved in Candida albicans biofilm persistence against miconazole[J].Antimicrob Agents Chemother,2011,55(9):4033-4037.
[17]Delattin N,Cammue BP,Thevissen K.Reactive oxygen species-inducing antifungal agents and their activity against fungal biofilms[J].Fut Med Chem,2014,6(1):77-90.