添加銀元素於鐵-22鉻-5鎳不銹鋼合金對抗菌性質及相關影響因子之研究

Researches of Antimicrobial Property and Related Influencing Factors on Fe-22Cr-5Ni Stainless Steels Alloy with Silver Elements

杜哲光(Je-Kang Du)

指導教授 ： 王兆祥

共同指導教授 ： 陳克恭(Ker-Kong Chen)

高雄醫學大學/口腔醫學院/牙醫學系博士班/博士(2017年)

摘要

不銹鋼與鈦金屬因抗腐蝕能力佳，且擁有不錯之機械性質，被廣泛應用於一般民生、工業設備及醫療器材之應用。在醫療應用上，常被使用的的合金為貴金屬、Ti-6Al-4V與316 L不銹鋼。AISI 2205雙相不銹鋼其具有Ti-6Al-4V的強度與316 L不銹鋼之彈性模數與延伸率，換言之，2205不銹鋼同時具有典型商用牙材之機械性質特性。同時，2205不銹鋼之價格亦具相當優勢。因此，2205不銹鋼，極具有生醫材料應用之潛力。
就抗菌能力而言，銀元素抗菌效果大於銅元素，但銀元素對人體所產生的毒性遠小於銅元素。2205雙相不銹鋼在醫療上，主要研究以表面塗銀抗菌性為主，且表面噴塗銀可達到抗菌效果；惟表面噴塗的銀含量較多，會使造成毒性產生進而對人體產生影響性。另外，表面處理會因使用產生磨損，而降低其功能。故考慮將2205不銹鋼製成塊材，並添加微量的銀元素分析銀元素在合金塊材中；經不同熱處理後，分析其機械性質、磨耗實驗與顯微結構的變化特徵。同時，探討合金之生物相容性(LDH與MTT測試)、抗菌性(E.coli與S.aureus測試)、電化學特性，以及動物活體驗證等評估。
鐵-22鉻-5鎳-x銀合金係以商用2205不銹鋼，添加0.1%、0.2%和0.3%銀，利用高週波冶煉爐進行精密鑄造。澆注溫度1650~1680℃，殼模溫度1250°C，鑄件尺寸為ψ=50.0±1mm，L=180.0±1 mm棒材，經1200°C均質化8小時後，於1050°C持溫30分鐘，自由模鍛成為t=3.0±0.05mm板材或ψ=15.0±0.5mm棒材；然後，CNC加工和線切割為研究所需之試片尺寸。
顯微結構分析顯示：微量的銀(Ag < 0.3%)對合金的相變態，沒有顯著的影響。合金經1080°C/30分鐘處理後，基本組織為(γ+42%α)雙相組織。在450°C處理，則可發現奈米級富銀相析出；提高溫度至650℃時，除奈米富銀相析出外，亦可觀察到X相析出物，其晶格常數a=0.261nm、c=0.468nm。另，高溫850℃與950℃處理時，亦可觀察到X相析出物，且發現M23C6的析出物沿著晶粒邊界析出，晶格常數a=1.028nm。機械性質分析顯示：銀的添加對合金的機械性質，沒有顯著的影響。合金經固溶處理後，其抗拉強度、降伏強度與延伸率，分別介於61.0~79.2kg/mm2、49.2~59.8kg/mm2與38.3~49.2%。高溫處理後，合金延伸率降至23.8~ 30.3%。顯示：富銀相析出對機械性質影響不大，X-phase與M23C6析出，則會影響合金之延伸率。
LDH與MTT(生物相容性)分析顯示：添加銀元素後，銀含量為0.3%時毒性值降低，測得之含量去氫酵素(dehydrogenase)較低，細胞死亡率也較低。E. coli與S. aureus (抗菌性)分析顯示：添加銀元素後，合金抗菌性提高，其中對E. coli抗菌特徵大於S. aureus。離子溶出分析顯示，抗菌性與銀離子溶出，呈現一致性，銀元素溶出越多，抗菌性越高，整體而言，銀的添加有助於合金之生物相容性與抗菌特徵表現，合金經固溶與低溫450 ℃熱處理8小時候後，呈現最佳之生物相容性與抗菌特徵。合金於人工唾液下，腐蝕電位為-0.275±0.015eV；其電化學腐蝕電位與目前商用鈦合金與不銹鋼牙材相當。動物實驗顯示：銀元素添加會降低菌落的組成，根據TGF-β1、caspase-3、LC3-II與IL-1β等分析實驗數據顯示：鐵-22鉻-5鎳-x銀合金具有抗菌能力，但過度的抗菌能力會破壞細胞，並抑制組織的自我修復功能，減緩細胞程序性死亡的能力。
整體而言，鐵-22鉻-5鎳-x銀合金之材料性質，不論在強度、延伸率與抗腐蝕性，極具牙科應用之潛力。合金之生醫特性，包括：細胞存活率與死亡率、E. coli與S.aureus抗菌性，以及動物實體植入分析，亦呈現優異的特性。

關鍵字

雙相不銹鋼 ； 機械性質 ； 顯微結構 ； 抗菌性 ； 塊材

並列摘要

According to well corrosion resistances and mechanical properties, the Titanium alloys and austenitic stainless steels are widely used in households, industrial and medical equipment. For medical applications, the typical alloys are the Ti–6Al–4V alloy and AISI 316L stainless steel. AISI 2205 duplex stainless steel possesses the strength similar to Ti–6Al–4V alloy and the modulus of elasticity and elongation similar to 316L stainless steel. In other words, 2205 stainless steel has the mechanical properties of typical commercial dental materials. Meanwhile, compared with the commercial medical material, the price of 2205 stainless steel is also a great advantage. Therefore, the 2205 stainless steel would be a potential application on the medical material, particularly on dental-orthodontic material.
As far as antibacterial ability is concerned, silver elements surpass copper, with the former’s toxicity being much lower. For the antibacterial ability and lower human’s toxicity, the silver element added on 2205 duplex stainless steel by surface coating is widely investigated by many workers, which the silver-coated surface could achieved antibacterial effects. Meanwhile, the coating film may wear with time resulting to decrease the antibacterial effects. Therefore, this study is focused on a 2205 bulk stainless steel with 0.1~0.3% silver element to prolong its antibacterial effects. In addition, the antibacterial effects and mechanical properties of the 2205 stainless steel with different microstructures are also investigated during various heated treatments. Meanwhile, the alloy biocompatibility (LDH and MTT tests), antibacterial properties (E. coli and S. aureus tests), and electrochemical characteristics were explored and animals were evaluated in vivo.
The ingots of present alloys (defined as Fe-22Cr-5Ni-xAg, x=0.1%, 0.2%, and 0.3%) were prepared by the commercial AISI 2205 stainless steel and 99.99% silver by using of air induction furnace, which the pouring temperature being 1650~1680°C and the cell temperature being 1250°C, respectively. The diameter of ingots are ?I=50.0±0.5mm and L=180.0±0.5mm. After being homogenization at 1200°C for 8 hours, and the ingots are hot-forged at 1050°C to be a t=3.0 ± 0.05mm plate or a ?I=15.0±0.5 mm bar. And then, the plate or bar are machining by using CNC machine and/or wire cutting machine to the specimens for series studies.
Based the analyses of microstructures, it reveals that the effects of silver added on the present alloys is no significant. According to SEM and TEM analyses, the microstructure of the as-quenched specimen is essential a (γ+42%α) duplex phase. When aged at 450°C, some Ag-rich nanoscale precipitates with a size of about 30 nm could be observed with the matrix. Prolonged the aging time and/or increasing the aging temperature to 650°C, the nanoscale precipitate would be loss coherence to be a semi-coherent precipitate. And, some plate-like x-phase precipitates are also found within ferrite matrix during 650°C aging processes. The lattice parameter of the x-phase precipitate is a=0.261nm and c=0.468nm. When aged at 850°C and 950°C, some x-phase precipitates could be also observed. In addition, some M23C6 carbides with a lattice parameter a=1.028nm could formed on grain boundaries. Based the analyses of mechanical properties, it also reveals that the effects of silver added on the present alloys is no significant. The tensile strength, yield strength and elongation of the present alloys with various heated treatment are in the range between 61.0~79.2kg/mm2, 49.2~59.8kg/mm2 and 38.3~49.2%, respectively. However, the elongation of the present alloy would be decreased to 23.8~30.3% during high-temperature annealing. It implied that the effects of nanoscale precipitates on the mechanical properties is no significant, and the precipitation of X-phase and M23C6 would decreased on the elongation.
Based on the LDH and MTT analyses, it shows that the addition silver element on the present alloy could reduce toxicity, with a smaller measured amount of dehydrogenase and a lower cell death rate. The examination of E. coli and S. aureus shows that the antibacterial properties of the present alloys were improved with silver element added. Meanwhile, the antibacterial characteristics of E. coli exhibited stronger than that of S. aureus. Moreover, the antibacterial characteristic of present alloys is consistence with the Ag+ dissolution. The optimal antibacterial characteristic would be found on the specimen aged at 450 °C for 8 hours. Anodic polarization at 0.03V/min in artificial saliva, the corrosion potential of the present alloys is near -0.275eV, of which is similar to the commercial dental material such as Titanium alloy and austenite stainless steel. According to animal experiments, it shows that the addition of silver element on the present alloys could reduce the formation of bacterial colonies. And, based on the analysis of TGF-β1, caspase-3, LC3-II and IL-1β, it also reveals that the Fe-22Cr-5Ni-0.2Ag alloy (Alloy C) possessed antibacterial ability. However, excessive antibacterial ability of Alloy C could damage cells and inhibit the self-healing function of organisms. And excessive antibacterial ability of alloy C could also slow down the capability of programmed cell death.
As mention above, the material properties of Fe-22Cr-5Ni-xAg such as strength, elongation and corrosion resistance will indicate a higher potential application on dental-orthodontic material. Based on the LDH, MTT, E. coli, S. aureus and animal studies, it is revealed that the bio-characteristics of present alloys are also excellent.

並列關鍵字

duplex stainless steel ； mechanical property ； microstructure ； antibacterial property ； bulk

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