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研究生： 楊舒雯
Yang, Shu-Wen
論文名稱： 赤蘚紅與金奈米粒子於光動力殺菌的協同作用之研究
Study of the Synergy Effect between Erythrosine and Gold Nanoparticles in Photodynamic Inactivation
指導教授： 施士塵
Shi, Shih-Chen
學位類別： 碩士
Master
系所名稱： 工學院 - 機械工程學系
Department of Mechanical Engineering
論文出版年： 2018
畢業學年度： 106
語文別： 中文
論文頁數： 87
中文關鍵詞： 光動力殺菌 、活性氧物種 、金奈米粒子 、協同作用 、單線態氧
外文關鍵詞： photodynamic inactivation, reactive oxygen species, gold nanoparticles, synergy effect, singlet oxygen
相關次數： 點閱：133  下載：4 
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    • 光動力治療(Photodynamic Therapy, PDT)是藉由光敏劑 (Photosensi-tizer, PS)暴露在特定波長的光源下，使光敏物質對細胞產生各種自由基 (Radicals)或單線態氧 (Singlet Oxygen, 1O2)等活性氧物種 (Reac-tive oxygen species, ROS)，並造成氧化性損傷。本實驗選用赤蘚紅 (Erythrosine)作為光敏劑，並搭配綠色LED光源 (525 nm)，針對大腸桿菌 (Escherichia coli, E. coli)進行光動力殺菌 (Photodynamic in-activation, PDI)測試，探討赤蘚紅搭配綠光的殺菌作用，再確認其殺菌效果後，考慮金奈米粒子良好的生物相容性且常與抗生素結合，具有攜帶及協助運送之功能，因此決定將其添加於實驗系統中，希望在不影響細菌生長情況下，比較兩者的殺菌效果。
    進行PDI後，藉由CFU(菌落形成單位)平板計數及Live/Dead細胞染色試劑確認利用赤蘚紅搭配綠色LED光源 (PS+, L+)，在短時間作用下，即可有良好的殺菌效果。同時亦發現在單獨使用金奈米粒子時，並不會影響細菌生長，但搭配光敏劑使用時 (PS+, Au+, L+)，有助於提升殺菌效率；藉由SEM觀測細菌表面形貌，發現金奈米粒子吸附於細菌表面，因此根據實驗結果推測金奈米粒子能夠攜帶光敏劑產生協同作用 (Synergy effect)，使得光敏劑能更準確地與細菌結合。為探討細菌死亡機制，進行ROS之檢測，並藉由添加不同的ROS消滅劑推測本實驗系統是因為單線態氧 (singlet oxygen)之產生 (Type II)，而導致細菌死亡。

    In the experiment, we did the photodynamic inactivation (PDI) test against E. coli by choosing erythrosine as a photosensitizer and using green LED lamps as the light source, and discussed the bactericidal action of the (PS+, L+) system. In the meanwhile, we considered the great biocompatibility of gold nanoparticles which have the function of assisting delivery. Therefore, we added the particles into the (PS+, Au+, L+) system and compared the bactericidal effect with the (PS+, L+) system. After PDI treatment, we did the colony-forming unit (CFU) counting and the LIVE/DEAD bacterial viability test. We demonstrated that E. coli could be in-activated by the (PS+, L+) system in a short time, and found that using gold nano-particles alone would not affect the growth of E. coli, but mixing with Erythrosine would increase the bactericidal efficiency. SEM images of E. coli with PDI treat-ments (PS+, Au+, L+) showed that gold nanoparticles would adhere to the surface of E. coli. According to the results, we inferred that gold nanoparticles could carry the photosensitizer to produce synergy effect, so that the photosensitizer can target bacteria precisely. Finally, in order to investigate the death mechanism of E. coli, we conducted the ROS test by adding different kinds of scavengers. We concluded that the death of E. coli was due to the production of singlet oxygen (Type II reac-tion) in our system.

    口試合格證明 I 摘要 II Extended abstract III 誌謝 XII 總目錄 XIV 表目錄 XVII 圖目錄 XVIII 第1章 緒論 1 1-1 前言 1 1-2 研究動機 3 第2章 文獻回顧 4 2-1 抗生素 4 2-2 奈米粒子之抗菌應用 8 2-3 金奈米粒子結合抗生素應用 13 2-4 光動力殺菌 (Photodynamic inactivation) 18 第3章 實驗內容 27 3-1 實驗目的 27 3-2 實驗簡介 27 3-3 實驗流程 28 3-4 實驗方法 29 3-4-1 細菌培養 29 3-4-2 O.D.值測量原理 29 3-4-3 光敏劑及光源選擇 29 3-4-4 金奈米粒子(Au NPs)之合成 32 3-4-5 金奈米粒子(Au NPs)之檢測 34 3-4-6 光動力殺菌(Photodynamic inactivation, PDI) 34 3-4-7 CFU平板計數 36 3-4-8 Live/Dead細菌死活檢測 37 3-4-9 SEM細菌表面形貌分析 40 3-4-10 活性氧物種(Reactive oxygen species, ROS)檢測 41 3-4-11 Type I及Type II 之檢測 42 3-5 實驗設備 44 第4章 實驗結果與討論 46 4-1 金奈米粒子之合成 46 4-1-1 金奈米粒子之顏色及吸收光譜檢測 46 4-1-2 金奈米粒子之粒徑分析 47 4-2 光動力殺菌(PDI) 51 4-2-1 CFU平板計數 51 4-2-1-1 赤蘚紅搭配綠色LED光源模組進行PDI 51 4-2-1-2 金奈米粒子對大腸桿菌之毒性測試 53 4-2-1-3 添加金奈米粒子進行PDI 55 4-2-2 Live/Dead細菌死活檢測 62 4-3 SEM-細菌表面形貌 68 4-4 金奈米粒子於PDI之作用機制推論 71 4-5 死亡機制討論 74 4-5-1 活性氧物種(ROS)檢測 74 4-5-2 Type I及Type II 之檢測 78 第5章 總結 81 5-1 實驗重點整理 81 5-2 結論 82 參考文獻 83

    [1] R. Ackroyd, C. Kelty, N. Brown, M. Reed,The history of photodetection and photodynamic therapy, Photochemistry and photobiology, 74 (2001) pp.656-669.
    [2] R.-M. Szeimies, J. Dräger, C. Abels, M. Landthaler, History of photodynamic therapy in dermatology, in: Comprehensive Series in Photosciences, Elsevier, (2001), pp. 3-15.
    [3] R.M. Amin, M.B. Mohamed, M.A. Ramadan, T. Verwanger, B. Krammer,Rapid and sensitive microplate assay for screening the effect of silver and gold nanoparticles on bacteria, Nanomedicine, 4 (2009) pp.637-643.
    [4] R. Gaynes,The Discovery of Penicillin—New Insights After More Than 75 Years of Clinical Use, Emerging Infectious Diseases, 23 (2017) pp.849-853.
    [5] H.C. Neu,The crisis in antibiotic resistance, Science, 257 (1992) pp.1064-1073.
    [6] M.C. McManus,Mechanisms of bacterial resistance to antimicrobial agents, American Journal of Health-System Pharmacy, 54 (1997) pp.1420-1433.
    [7] K. Drlica, X. Zhao,DNA gyrase, topoisomerase IV, and the 4-quinolones, Microbiology and molecular biology reviews, 61 (1997) pp.377-392.
    [8] F.C. Tenover,Mechanisms of antimicrobial resistance in bacteria, American journal of infection control, 34 (2006) pp.S3-S10.
    [9] D.R. Storm, K.S. Rosenthal, P.E. Swanson,Polymyxin and related peptide antibiotics, Annual review of biochemistry, 46 (1977) pp.723-763.
    [10] D. Brown,Antibiotic resistance breakers: can repurposed drugs fill the antibiotic discovery void?, Nature Reviews Drug Discovery, 14 (2015) pp.821.
    [11] A.H. Holmes, L.S. Moore, A. Sundsfjord, M. Steinbakk, S. Regmi, A. Karkey, P.J. Guerin, L.J. Piddock,Understanding the mechanisms and drivers of antimicrobial resistance, The Lancet, 387 (2016) pp.176-187.
    [12] I. Sondi, B. Salopek-Sondi,Silver nanoparticles as antimicrobial agent: a case study on E. coli as a model for Gram-negative bacteria, Journal of colloid and interface science, 275 (2004) pp.177-182.
    [13] Q.L. Feng, J. Wu, G. Chen, F. Cui, T. Kim, J. Kim,A mechanistic study of the antibacterial effect of silver ions on Escherichia coli and Staphylococcus aureus, Journal of biomedical materials research, 52 (2000) pp.662-668.
    [14] Y. Liu, L. He, A. Mustapha, H. Li, Z. Hu, M. Lin,Antibacterial activities of zinc oxide nanoparticles against Escherichia coli O157: H7, Journal of applied microbiology, 107 (2009) pp.1193-1201.
    [15] J.W. Chan, H. Winhold, M.H. Corzett, J.M. Ulloa, M. Cosman, R. Balhorn, T. Huser,Monitoring dynamic protein expression in living E. coli. Bacterial cells by laser tweezers Raman spectroscopy, Cytometry Part A, 71 (2007) pp.468-474.
    [16] P. Gajjar, B. Pettee, D.W. Britt, W. Huang, W.P. Johnson, A.J. Anderson,Antimicrobial activities of commercial nanoparticles against an environmental soil microbe, Pseudomonas putida KT2440, Journal of Biological Engineering, 3 (2009) pp.9.
    [17] N. Kumar, V. Shah, V.K. Walker,Perturbation of an arctic soil microbial community by metal nanoparticles, Journal of hazardous materials, 190 (2011) pp.816-822.
    [18] Y. Zhang, T.P. Shareena Dasari, H. Deng, H. Yu,Antimicrobial activity of gold nanoparticles and ionic gold, Journal of Environmental Science and Health, Part C, 33 (2015) pp.286-327.
    [19] I.A. Wani, T. Ahmad,Size and shape dependant antifungal activity of gold nanoparticles: a case study of Candida, Colloids and surfaces B: Biointerfaces, 101 (2013) pp.162-170.
    [20] C. Jayaseelan, R. Ramkumar, A.A. Rahuman, P. Perumal,Green synthesis of gold nanoparticles using seed aqueous extract of Abelmoschus esculentus and its antifungal activity, Industrial Crops and Products, 45 (2013) pp.423-429.
    [21] S. Shankar, L. Jaiswal, R. Aparna, R. Prasad,Synthesis, characterization, in vitro biocompatibility, and antimicrobial activity of gold, silver and gold silver alloy nanoparticles prepared from Lansium domesticum fruit peel extract, Materials Letters, 137 (2014) pp.75-78.
    [22] J.F. Hernández-Sierra, F. Ruiz, D.C.C. Pena, F. Martínez-Gutiérrez, A.E. Martínez, A.d.J.P. Guillén, H. Tapia-Pérez, G.M. Castañón,The antimicrobial sensitivity of Streptococcus mutans to nanoparticles of silver, zinc oxide, and gold, Nanomedicine: Nanotechnology, Biology and Medicine, 4 (2008) pp.237-240.
    [23] A. Azam, F. Ahmed, N. Arshi, M. Chaman, A. Naqvi,One step synthesis and characterization of gold nanoparticles and their antibacterial activities against E. coli (ATCC 25922 strain), Int J Theor Appl Sci, 1 (2009) pp.1-4.
    [24] V.D. Badwaik, L.M. Vangala, D.S. Pender, C.B. Willis, Z.P. Aguilar, M.S. Gonzalez, R. Paripelly, R. Dakshinamurthy,Size-dependent antimicrobial properties of sugar-encapsulated gold nanoparticles synthesized by a green method, Nanoscale research letters, 7 (2012) pp.623.
    [25] Y. Zhou, Y. Kong, S. Kundu, J.D. Cirillo, H. Liang,Antibacterial activities of gold and silver nanoparticles against Escherichia coli and bacillus Calmette-Guérin, Journal of nanobiotechnology, 10 (2012) pp.19.
    [26] H. Gu, P. Ho, E. Tong, L. Wang, B. Xu,Presenting vancomycin on nanoparticles to enhance antimicrobial activities, Nano letters, 3 (2003) pp.1261-1263.
    [27] B. Saha, J. Bhattacharya, A. Mukherjee, A. Ghosh, C. Santra, A.K. Dasgupta, P. Karmakar,In vitro structural and functional evaluation of gold nanoparticles conjugated antibiotics, Nanoscale research letters, 2 (2007) pp.614.
    [28] A.N. Grace, K. Pandian,Antibacterial efficacy of aminoglycosidic antibiotics protected gold nanoparticles—A brief study, Colloids and Surfaces A: Physicochemical and Engineering Aspects, 297 (2007) pp.63-70.
    [29] M. Chamundeeswari, S. Sobhana, J.P. Jacob, M.G. Kumar, M.P. Devi, T.P. Sastry, A.B. Mandal,Preparation, characterization and evaluation of a biopolymeric gold nanocomposite with antimicrobial activity, Biotechnology and applied biochemistry, 55 (2010) pp.29-35.
    [30] D. Pornpattananangkul, L. Zhang, S. Olson, S. Aryal, M. Obonyo, K. Vecchio, C.-M. Huang, L. Zhang,Bacterial toxin-triggered drug release from gold nanoparticle-stabilized liposomes for the treatment of bacterial infection, Journal of the American Chemical Society, 133 (2011) pp.4132-4139.
    [31] A.N. Brown, K. Smith, T.A. Samuels, J. Lu, S.O. Obare, M.E. Scott,Nanoparticles functionalized with ampicillin destroy multiple-antibiotic-resistant isolates of Pseudomonas aeruginosa and Enterobacter aerogenes and methicillin-resistant Staphylococcus aureus, Applied and environmental microbiology, 78 (2012) pp.2768-2774.
    [32] B. Nagaraj, T. Divya, B. Malakar, N. Krishnamurthy, R. Dinesh, C. Negrila, C. Ciobanu, S. Iconaru,Phytosynthesis of gold nanoparticles using Caesalpinia pulcherrima (peacock flower) flower extract and evaluation of their antimicrobial activities, Dig J Nanomater Biostruct, 7 (2012) pp.899-905.
    [33] L. Costa, M.A. Faustino, J.P. Tomé, M.G. Neves, A.C. Tomé, J.A. Cavaleiro, Â. Cunha, A. Almeida,Involvement of type I and type II mechanisms on the photoinactivation of non-enveloped DNA and RNA bacteriophages, Journal of Photochemistry and Photobiology B: Biology, 120 (2013) pp.10-16.
    [34] A.B. Ormond, H.S. Freeman,Dye sensitizers for photodynamic therapy, Materials, 6 (2013) pp.817-840.
    [35] C. Chui, K. Hiratsuka, A. Aoki, Y. Takeuchi, Y. Abiko, Y. Izumi,Blue LED inhibits the growth of Porphyromonas gingivalis by suppressing the expression of genes associated with DNA replication and cell division, Lasers in surgery and medicine, 44 (2012) pp.856-864.
    [36] Y.-H. Lee, H.-W. Park, J.-H. Lee, H.-W. Seo, S.-Y. Lee,The photodynamic therapy on Streptococcus mutans biofilms using erythrosine and dental halogen curing unit, International journal of oral science, 4 (2012) pp.196.
    [37] L.P. Rosa, F.C. da Silva, S.A. Nader, G.A. Meira, M.S. Viana,Antimicrobial photodynamic inactivation of Staphylococcus aureus biofilms in bone specimens using methylene blue, toluidine blue ortho and malachite green: an in vitro study, Archives of oral biology, 60 (2015) pp.675-680.
    [38] B. Zeina, J. Greenman, W. Purcell, B. Das,Killing of cutaneous microbial species by photodynamic therapy, British Journal of dermatology, 144 (2001) pp.274-278.
    [39] S.-C. Shi, W.-K. Huang,Evaluation of Photodynamic Inactivation Efficiency Using Conventional and Decorative Light-Emitting Diode Lamps, Sensors and Materials, 29 (2017) pp.1569-1577.
    [40] S.F.G. Vilela, J.C. Junqueira, J.O. Barbosa, M. Majewski, E. Munin, A.O.C. Jorge,Photodynamic inactivation of Staphylococcus aureus and Escherichia coli biofilms by malachite green and phenothiazine dyes: an in vitro study, Archives of oral biology, 57 (2012) pp.704-710.
    [41] J.-H. Park, Y.-H. Moon, I.-S. Bang, Y.-C. Kim, S.-A. Kim, S.-G. Ahn, J.-H. Yoon,Antimicrobial effect of photodynamic therapy using a highly pure chlorin e6, Lasers in medical science, 25 (2010) pp.705-710.
    [42] A. Pfitzner, B.W. Sigusch, V. Albrecht, E. Glockmann,Killing of periodontopathogenic bacteria by photodynamic therapy, Journal of periodontology, 75 (2004) pp.1343-1349.
    [43] R.D. Rossoni, J.C. Junqueira, E.L.S. Santos, A.C.B. Costa, A.O.C. Jorge,Comparison of the efficacy of Rose Bengal and erythrosin in photodynamic therapy against Enterobacteriaceae, Lasers in medical science, 25 (2010) pp.581-586.
    [44] A.C.B.P. Costa, V.M. de Campos Rasteiro, C.A. Pereira, E.S.H. da Silva Hashimoto, M.B. Junior, J.C. Junqueira, A.O.C. Jorge,Susceptibility of Candida albicans and Candida dubliniensis to erythrosine-and LED-mediated photodynamic therapy, Archives of oral biology, 56 (2011) pp.1299-1305.
    [45] D. Min, J. Boff,Chemistry and reaction of singlet oxygen in foods, Comprehensive reviews in food science and food safety, 1 (2002) pp.58-72.
    [46] D.B. Min, H.-O. Lee, Chemistry of lipid oxidation, in: Flavor Chemistry, Springer, (1999), pp. 175-187.
    [47] N. Kashef, Y.-Y. Huang, M.R. Hamblin,Advances in antimicrobial photodynamic inactivation at the nanoscale, Nanophotonics, 6 (2017) pp.853-879.
    [48] K. Gollnick,Type II photooxygenation reactions in solution, Advances in Photochemistry, Volume 6, (2007) pp.1-122.
    [49] L. Huang, Y. Xuan, Y. Koide, T. Zhiyentayev, M. Tanaka, M.R. Hamblin,Type I and Type II mechanisms of antimicrobial photodynamic therapy: An in vitro study on gram‐negative and gram‐positive bacteria, Lasers in surgery and medicine, 44 (2012) pp.490-499.
    [50] C.S. Foote, Type I and type II mechanisms of photodynamic action, in, ACS Publications, (1987).
    [51] D.E. Dolmans, D. Fukumura, R.K. Jain,Photodynamic therapy for cancer, Nature reviews cancer, 3 (2003) pp.380.
    [52] J.-Y. Liang, J.-M.P. Yuann, C.-W. Cheng, H.-L. Jian, C.-C. Lin, L.-Y. Chen,Blue light induced free radicals from riboflavin on E. coli DNA damage, Journal of Photochemistry and Photobiology B: Biology, 119 (2013) pp.60-64.
    [53] M. Salmon-Divon, Y. Nitzan, Z. Malik,Mechanistic aspects of Escherichia coli photodynamic inactivation by cationic tetra-meso (N-methylpyridyl) porphine, Photochemical & Photobiological Sciences, 3 (2004) pp.423-429.
    [54] X. Ji, X. Song, J. Li, Y. Bai, W. Yang, X. Peng,Size control of gold nanocrystals in citrate reduction: the third role of citrate, Journal of the American Chemical Society, 129 (2007) pp.13939-13948.
    [55] A. Tavares, S.R. Dias, C.M. Carvalho, M.A. Faustino, J.P. Tomé, M.G. Neves, A.C. Tomé, J.A. Cavaleiro, Â. Cunha, N.C. Gomes,Mechanisms of photodynamic inactivation of a Gram-negative recombinant bioluminescent bacterium by cationic porphyrins, Photochemical & Photobiological Sciences, 10 (2011) pp.1659-1669.
    [56] R.S. Bodannes, P.C. Chan,Ascorbic acid as a scavenger of singlet oxygen, Febs Letters, 105 (1979) pp.195-196.
    [57] H. Fukumura, M. Sato, K. Kezuka, I. Sato, X. Feng, S. Okumura, T. Fujita, U. Yokoyama, H. Eguchi, Y. Ishikawa,Effect of ascorbic acid on reactive oxygen species production in chemotherapy and hyperthermia in prostate cancer cells, The Journal of Physiological Sciences, 62 (2012) pp.251-257.
    [58] S. Eustis, M.A. El-Sayed,Why gold nanoparticles are more precious than pretty gold: noble metal surface plasmon resonance and its enhancement of the radiative and nonradiative properties of nanocrystals of different shapes, Chemical society reviews, 35 (2006) pp.209-217.
    [59] S.K. Ghosh, T. Pal,Interparticle coupling effect on the surface plasmon resonance of gold nanoparticles: from theory to applications, Chemical reviews, 107 (2007) pp.4797-4862.

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