Characterization of the silver thin films produced at different substrate temperatures

Science & Technology Development Journal, 22(4):356- 364 Open Access Full Text Article Original Research 1Faculty of Physics and Engineering Physics, VNUHCM-University of Science 2Falculity of Physics, Dong Thap University 3Faculty of Chemistry, VNUHCM-University of Science Correspondence Vu Thi Hanh Thu, Faculty of Physics and Engineering Physics, VNUHCM-University of Science Email: vththu@hcmus.edu.vn History  Received: 2019-06-30  Accepted: 2019-11-14  Published: 2019-12-31 DOI : 10.32508/stdj.v22i4.1691 Copyright © VNU-HCM Press. This is an open- access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Characterization of the silver thin films produced at different substrate temperatures Ton Nu Quynh Trang1, Le Thi Ngoc Tu2, Tran VanMan3, Vu Thi Hanh Thu1,* Use your smartphone to scan this QR code and download this article ABSTRACT Introduction: In recent decades, the antimicrobial surfaces/coating properties towards a long- lasting microbicidal effect have drawn enormous attention by researchers, they have been devel- oped and used in a wide variety of high-touch hospital devices as a potential approach. Methods: In this work, Ag NPs was synthesized by sputtering method at the different annealing tempera- tures of 100C, 200C, 300C, and 400C. Results: As a result, the as-synthesized Ag-300 exhibits the highest E. coli antibacterial performance compared with others. This can be attributed to the change of the Ag NPs toxicity based on the growth of nanoparticles during the deposition process related to the Ostwald ripening process, thermal activation and coalescence particles. Conclu- sion: This work provides an essential insight into the antimicrobial activity of Ag NPs-based films synthesized through the vacuum deposition technique, resulting in opening a new approach for enhancing the antimicrobial efficacy and prospects. Keywords: Silver nanoparticles, antibacterial mechanism, vacuum deposition techniques, toxicity INTRODUCTION Nanoparticles (NPs) have been considered as one of the most promising alternatives for traditional ma- terials in many fields of science and technology 1,2, due to nanoscale approaching physical characteris- tics and functionalities that can be assigned to the different from their bulk counterparts. As an exam- ple, the antibacterial performance of nanomaterials such as ZnO, TiO2, and Ag NPs have drawn enor- mous attention in recent years for their desired appli- cations in biomedical applications, water disinfection, and consumer goods3,4. However, a current draw- back is the synthesis and application of nanoparti- cles to implement effective measures that can prevent wound infections burns and chronic ulcers associated with healthcare caused by microorganisms. Specifi- cally, healthcare-associated infections have been con- sidered as one of the global threats related to bacterial pathogen emergence and one of the main reasons for patient morbidity and mortality. Indeed, an approxi- mated 20% to 40% of health associated with the infec- tion fields was reported 5. Among all nanomaterials, silver nanoparticles (Ag NPs) not only have attracted more attention in healthcare-associated fields but also considered as one of the most promising candidates for potential medical applications in recent years due to their unique nano-physicochemical characteristics and broad-spectrum antimicrobial activity 6,7. Previ- ous studies reported that Ag NPs, which synthesized in a similar way to form silver ions, exerted against bacteria through a multifactorial process, and they were associated with inhibiting the growth of harmful bacteria as they were harm to the bacterial cell wall and plasma membrane or restraint on DNA replica- tion and protein8. They could be released by the nat- ural formation of ion Ag in the presence of reductive components in the environment9. Moreover, the characteristic features of nanoparticles such as size, shape, density distribution have been demonstrated to affect the antibacterial activity of Ag NPs significantly, this could be attributed to a dif- ferential release of Ag+ ions10. Previous researches in this field suggested that the effects of shapes and sizes prepared by wet chemical reduction methods or biosynthesis play a significant role in the antimicro- bial nature of Ag NPs11–14. These methods have been devoted to controlling the size and shape of the Ag NPs, however, still, some obstacles such as the pres- ence of colloidal stabilizers or impurities, the toxic solvents, and the sophisticated synthesis process to re- duce or suppress the aggregation phenomenon in so- lution were detected15–17. Good adhesion between the AgNPs and the substrate has great potential in the practical applications, while, themere synthesis of the Ag NPs is rather hard for most of the desired appli- cations. Therefore, an explored alternative route has been centered on solving the aforementioned draw- backs of wet chemical methods. Amongst, vacuum Cite this article : Quynh Trang T N, Ngoc Tu L T, Van Man T, Hanh Thu V T. Characterization of the silver thin films produced at different substrate temperatures. Sci. Tech. Dev. J.; 22(4):356-364. 356 Science & Technology Development Journal, 22(4):356-364 deposition techniques consist of thermal evapora- tion, magnetron sputtering and pulsed laser deposi- tion have drawn enormous attention in recent years and considered as an effectivemethod to expand their applications because it can be grown on large surface areas with good quality, easy control in fabrication process, and environmentally friendly based on dif- ferent preparation conditions. In which, the temper- ature factor has been considered as one of the most critical factors for a reaction, adhesion and phase sep- arated morphology at the interface that can affect the change inmorphology on flat substrates in the growth process of the thin film significantly. Compared with other Ag types (Ag suspension, powder), Ag thin film on the flat substrates has some advantages i) they can save the material that can obtain the surface an- tibacterial performance equally ; ii) the diffusion of Ag nanoparticles into the environment is limited due to the good adhesion of Ag nanoparticles on the sub- strates deposited via sputtering method. In this work, Ag NP films have been prepared by DC magnetron sputtering directly on the surface of glass at the different annealing temperature from 100C to 400C to investigate the change of the crystalline structure, optical, morphology, and their antibacte- rial activity against various E. Coli bacteria. These results support the new approach for the design and synthesis of other precious metal as Ag thin films on plane substrates. METHODS Materials Ag target (with a purity of 99.99% and size of 76 x 5 mm, Advantage, Singapore), hydrochloric acid (HCl, 36%, Sigma-Aldrich), acetone (Sigma-Aldrich), dou- ble distilled water. All other chemicals were used as received without further purification. Preparation of Ag NPs film In this study, Ag nanostructured films were deposited at room temperature by using DC magnetron sput- tering on the corning glass (size of 76 x 26 x 1 mm, Marienfeld, Germany 900), and the base pressure was around 5 x105 Torr. First, the glass substrates were cleaned thoroughly in hydrochloric acid, ace- tone, then ultrasonicated in double distilled water in 15 min. Finally, substrates were dried under a stream of nitrogen. Before sputtering, argon plasmawas used to etch the surface of the substrate for 10 min in order to remove residual particles on the substrate surface. The substrate holder was rotated at a speed of 5 rpm during deposition. While the target was pre-sputtered for 5 min to dismiss contaminants and oxidized lay- ers. TheAg target power was set at 9W, the Ar gas was usedwith a flow rate of 18 sccm, and the total pressure was approximately 2.5 x 103 Torr. The amount of Ag was deposited with a constant time at 30 seconds, and the substrate temperatures are changed between 100C and 400C. Moreover, the corresponding sam- ples were denoted as Ag-y, where y was substrate tem- peratures. Characterization The structure and crystallinity of films were further investigated using X-ray diffraction measurements recorded using a Bruker D8 ADVANCE system with CuKa radiation source (l=0.154056 nm). The surface morphology of the films was observed using Hitachi S-4800 scanning electron microscopy (SEM, Hitachi S-4800) at room temperature, and atomic force mi- croscopy (AFM, SPM 5500, USA). The optical prop- erties of the films were characterized using a JASCO- V670 spectrophotometer ranging from 300 to 800 nm scan rate of 200 nm.min1 at room temperature. E. coli a ntibacterial experiment Bacterial culturing and plating were conducted fol- lowing the standard methods described in previous research18,19. The 1  106 colony-forming unit cul- ture was allowed to drip on both o n the surface of Corning glass containing the Ag NP film and in an unmodified glass slide (blank slide). They were then placed at room temperature. In the E.coli antibacterial performance test, all the experiments were conducted in a sterile environment. For all samples, the serial di- lutionwas done, and the dilutionwas then spread uni- formly on the surface into culture nutrient agar plates, and this plate was incubated at 37oC for 24 hours. The bactericidal activity of Ag NPs was investigated by the spread plate method. Finally, the number of colonies grown on the agar plates was counted and killing (%) efficacy of Ag NPs was calculated using the following equation: E.coli a ntibacterial efficacy (%) = [( Ncontrol – Ntreated)/Ncontrol ] x 100, where Ncontrol , Ntreated are numbers of bacteria grown on the agar plates follow- ing treatment with glass and Ag NPs films, respec- tively. RESULTS The crystal structures of the Ag NPs films were char- acterized by X-ray diffraction patterns, and the re- sults are shown in Figure 1. The diffraction peak ap- pears at 2 q = 38.2 and 44.4 corresponding to lat- 357 Science & Technology Development Journal, 22(4):356-364 tice plane (111), (200) of silver crystal particle, respec- tively (JCPDS cards no 04-0783). As shown in Fig- ure 1, t he XRD patterns are significantly changed at various substrate temperatures from 100C to 400C. It is clearly observed that the peak intensity of crys- tal planes further enhances with increasing substrate temperature, which is mainly governed by improving in crystallinity of the face-c entered cubic phase of Ag. The preferred orientation of the Ag along with (111) and (200) plane is observed. Also, these peaks become more dominant at higher substrate temperatures; this might be associated with the thermodynamic phase boundary and surface interdiffusion phenomenon. While surface interdiffusion considered as the dom- inant kinetic process that plays a central role in con- trolling the crystalline morphology of the film. No typical diffraction peaks corresponding to silver ox- ide are observed, which maybe below the XRD de- tection limit20. Moreover, the (111) plane has a high atomic density of electrons that hasmore favorable for highly reactive21. Therefore, their antibacterial effi- ciency againstE. Coli ofAgNPs is enhanced due to the interaction of the bacterial surface morphology with (111) plane. The prepared samples are further evaluated by SEM to reveal the morphology characteristics of Ag NPs, and the results are shown in Figure 2. As can be seen in Figure 2, when the substrate tem- perature increases, the particle size of Ag films in- creases leading to an increase in the mobility of sil- ver atoms on the surface. This can be indicated that the distribution of Ag atoms is extremely in homoge- neous that is governed by both local thermal energy and partially crystallizes the silver atoms. The par- ticle size increases from 10 to 50 for substrate tem- perature at 100 to 400, respectively. Among all these types, the Ag-400 sample tends to the coalesce and form clusters, an enlargement of the clusters is linked to each other forming a film with a large open area fraction. With increasing the substrate temperature, surface characteristics such as number density, shape, size, inter-particle distance, and surface were adjusted through some scenarios22,23. At room temperature of growth, the nanoparticles are almost disordered that may be attributed to the streams of atoms based on moving and collision with each other on the surface. After a further 100C, atom movement is faster, this has resulted in the formation of single crystalline. Af- ter 200C and above, the integration of atom clus- ters is almost completed with the rearrangement pro- cess during their progression to becoming more crys- talline. Moreover, the integration of many atom clusters into one governing structure can have a severe impact on the final structure of nanoparticles. This is attributed to the thermodynamic driving force that plays a cen- tral role in phase separation. However, the mecha- nism of phase separation is rather difficult to iden- tify continuous or discontinuous transformation for these thin films, which was deposited at an elevated temperature. These have resulted in frozen in place by the incoming flux as reported by Adams et al.24. That can highly impact the antibacterial activity of Ag nanoparticles. It is also interesting to consider whether Ostwald ripening processes, which aremainly governed by dif- fusion or coalescence of single atoms driven by a gra- dient in chemical potential causing the interchange of the neighboring atoms. This may also provide valu- able insight into the growth of larger nanoparticle, more stable particles. As regards, the neighboring of twoAgNPs (themarked yellow circles denoted as Par- ticle 1 and Particle 2) shows in Figure 2. Particle 1 re- duces in size, whereas, particle 2 achieves a larger size after increasing temperature deposition for 400C, in- dicating that the particle appearance through an Ost- wald ripening mechanism in this scenario ha s hap- pened that forms a dense close-packed structure for 400C as shown Figure 2d. These results indicate that this phenomenon plays a key role in the E. coli anti bacterial of Ag NPs films. In order to understand further the role of increasing substrate temperature to the evolution of surfacemor- phology, the root means square (RMS) roughness in Ag thin films was investigated by AFM analysis, as shown inFigures 3 and4. The results have shown that the particle size increases with increasing substrate temperature. This can be attributed to the increased mobility of silver atoms due to the arising of local ther- mal energy lead to the formation of the disordered Ag phase25. Moreover, the root means square and aver- age grain size of Ag thin films increase with arising the substrate temperature, as shown in Figure 4e. The RMS values of Ag thin films at various substrate tem- peratures were 0.8, 1.5, 2.2, 3.5, and 6.0 mm, respec- tively. Moreover, the surface appears rougher surface with more wrinkles (Figure 3d). Obviously, with in- creasing the substrate temperature from 100 to 400, the particle size and the RMS values increase, this is mainly attributed to the Ostwald ripening processes and the increment of energy surface at high tempera- ture. In the case of Ostwald ripening, the higher sur- face to volume ratio and formation of bigger particles appeared due to coalescence. These are increased the particle size and inter particle distance26. In order to 358 Science & Technology Development Journal, 22(4):356-364 Figure 1: XRD patterns of Ag thin films at different substrate temperatures from 100Cto 400C. Figure 2: SEM images of substrate temperature of Ag NPs films at (a) 100C, (b) 200C, (c) 300C, and (d) 400C, respectively. 359 Science & Technology Development Journal, 22(4):356-364 better understand the mechanism of the growth Ag with increasing substrate temperature, a schematic of the growth in morphology is illustrated in Figure 5. In order to provide further information about its im- pact on optical properties based on their stage ofmor- phological evolution, theUV-Visible absorption spec- troscopy was used to evaluate the optical properties of samples. The absorption spectra of Ag NPs films at different substrate temperatures are clearly displayed in Figure 6. The variation of distribution in parti- cle sizes of the Ag NPs film is attributed to the in- homogeneous broadening of the plasmon resonance peak. According toMie theory18, the particle size de- creased, the efficiency of absorption would be dom- inated over the scattering efficiency. Therefore, par- ticles with a small size would be given rise to en- hanced plasmon resonance. However, the distance between particles is large, so that coherent phase re- lations among the scattered light from different par- ticles was not observed. Moreover, the red shift of the peak in the dipole resonance has been observed with increasing the particle size, which is assigned to the weakening of the restoring force. This can be ex- plained by the increase of distance between charges on opposite sides of the particles, leading to a reduction in interaction. It is observed that the surface plasmon resonance (SPR) peak for Ag NPs films has a strong absorption at 450 nm corresponding to the dipolar resonance of Ag NPs. According to Figure 6 revealed that the AgNPs film at various substrate temperatures shows a broadening and red shift of SPR spectra. At substrate temperatures 100C, the dramatic broad- ening along with pronounced red shift is obtained. This broadening can be attributed to the increase in particle size due to not much change in surface cov- erage, except crystalline form can be transformed. As a result, the optical properties of Ag nanostructures were dependent on their surfacemorphology through their growth stages. It is well-known that silver has been considered as one of the most promising candidates to prevent infections for thousands of years. However, the silver-induced bactericidal effect is a complicated re- sponse related to the disruption of bacterial physiolo- gies, such as the formation of disulfide bonds, iron metabolism, and homeostasis. So, in order to fur- ther evaluate the relationship between the particle size and toxicity of Ag NPs in eradicating Escherichia coli pathogenic, the live bacteria cells are treated with Ag NPs films at different substrate temperatures 100, 200, 300, and 400C respectively, as delineated Figure 7. These results indicate that the percentage of surviv- ing bacteria reduces with increasing substrate tem- perature, suggesting that the antimicrobial efficacy in- creases while at 400C, the antimicrobial efficacy de- creases. The substrate temperature plays a vital role in the E. coli antibacterial activity that is assigned to the significant change in the particle size and surfacemor- phology of AgNPs, as displayed in Figures 2, 3 and 4. In which, Ag-300 exhibits the highest the E. coli an- timicrobial efficiency compared with others due to i) the high interaction between Ag NPs and E.coli bac- teria based on the uniform distribution between the adjacent AgNPs, ii) the high toxicity path of Ag NPs based on the regular shapes and appropriate sizes of Ag NPS. DISCUSSION It is a fact that the increase of Ag NPs particle size is significantly reduced the specific surface area. There- fore, the release of dissolvedAg species ismarkedly re- tarded. As a result, their toxicity responses are lower. Therefore, it is essential to investigate the change in the particle size that may be caused by the dramatic change in toxicity and surface area of Ag NPs. Recent studies have reported that the highly efficient E. coli antibacterial activity of Ag NPs can be ascribed to the large surface area due to the interactions between the surface area of particles and microbial cells. For instance, Morones et al. reported that the highly efficient antimicrobial performance of Ag NPs is due to the large surface area, which means that the per- centage of the interaction of small particles was higher than larger particles of the same parent material 19. Acharya et al. demonstrated that the distortion of the bacterial cell membrane byAg nanosphere was higher than one-dimension nanostructure (Ag nanorod), which could mainly be governed by the granulate shape with a larger specific surface area as compared with 1-Dimension nanostructure (Ag nanorod) with low specific surface area 27. Also, Hong et al. revealed that the nanosphere had a larger effective specific con- tact area, which means that they quickly achieved the interaction with bacterial cells and caused more dam- ages28. Thus, based on the above observations, it can be concluded that the specific surface area of Ag NPs plays a vital role in antimicrobial efficacy. Besides, it is clearly observed that AgNPswith small particle size is shown to bemore damaged as comparedwith larger particles in the silver-induced bactericidal because the smaller Ag NPS can be more easily penetrated E. coli bacteria than largerAgNPs orAgNPs aggregates. This also substantiate s the difference in the respective par- ticle size as shown in SEM images (Figure 2). Accord- ing to recent studies confirmed that the accumulation 360 Science & Technology Development Journal, 22(4):356-364 Figure 3: 2D AFM images of Ag thin films at various of substrate temperature (a) room temperature, (b) 100C, (c) 200C, (d) 300C, and (e) 400C. Figure 4: 3D AFM images of Ag thin film at various of substrate temperature (a) 100C, (b) 200C, (c) 300C, and (d) 400C. (e) RMS values and average grain size of Ag thin films at different substrate temperatures. 361 Science & Technology Development Journal, 22(4):356-364 Figure 5: Schematic of the growth inmorphology as a result of increasing substrate temperature. Figure 6: UV-vis spectra of different annealing temperatures from 100C to 400C of Ag NPs films. of different sizes of Ag NPs in the food chain was in- vestigated the toxicity of AgNPs and Ag+ ions during the antimicrobial process. The results revealed that some of the small Ag NPs remained in the digestive lumen, subcutaneous tissue, and gonad. This veri- fied that the Ag NPs uptake at the intracellular level and only small particles or partial aggregation of sil- ver nanoparticles were detected in the cells, whereas larger aggregation was not internalized in E. coli29–31. On the other hand, the antibacterial efficacy of Ag NPs films is affected by controlling the toxicity path of AgNPs based on changing the particle size. The small AgNPs inducedmore cellular toxicity than larger par- ticles. For example, Liu et al. reported that Ag NPs with size particle of 5 nm had more toxicity than 20 and 50 nm Ag NPs towards human cells32. Wang et al. reported that Ag NPs nanoparticle size of 10 nm was much more cytotoxic than 40 and 75 nm Ag NPs to human lung cells. However, toxicity between 10 nm citrate and 10 nmPVP-coatedAgNPswas not ob- served33. These results are well justified deduce that the Ag NPs toxicity related to their antimicrobial effi- ciency not only dependent particle size but also based on bacterial type. The toxicity plays a significant role in the antimicrobial capability of Ag NPs, which is at- tributed to the release of Ag+ cations that can be in- teracted with cells and intracellular macromolecules such as proteins and DNA. Besides, an accumulation of intercellular reactive oxygen species (ROS) reacts directly with protein and causes oxidative stress. So, the partial or permanent loss of structure and/or func- tion of the cellular proteinmay be caused by the above processes leads to the bacterial growth is inhibited. On the other side, the Ag NPs shows excellent E. coli antibacterial effect at different substrate temperature due to the presence of (111) plane. This can be at- tributed to the plane contains a high atomic density of electrons. Overall, our results showcased that Ag NPs films can inhibit the growth of bacteria based on the density distribution and particle size of Ag NPs leading to the change of their toxicity. CONCLUSIONS In summary, we have investigated the antimicrobial activity of Ag NPs films prepared by DC magnetron sputtering technique. Their optical properties were analyzed by UV-Vis spectroscopy, while, the struc- ture and morphological properties were investigated by XRD, FE-SEM, and AFM, respectively. Besides, 362 Science & Technology Development Journal, 22(4):356-364 Figure 7: Development of E. coli colonies (a) and E. coli antibacterial activity of different substrate temperatures of Ag NPs films against E. coli calculated by plate count method (b). their antimicrobial efficacy is tuned by adjusting the change of morphology of Ag NPs, such as density distribution and particle size. The results show that the E. coli anti-bacterial efficacy of the Ag-300 sample could reach up to~100%. This can be attributed to the change in the particle size and surface morphology of Ag NPs leading to the change of the Ag NPs toxicity. The work also provides a better understanding of the effect of Ag NPs morphology in enhancing the sur- face plasmon resonance related to Ostwald ripening processes that can be developedmore efficient desired antimicrobial systems under the visible regime in the future. ABBREVIATIONS NPs: Nanoparticles Ag NPs: Silver nanoparticles COMPETING INTERESTS The authors declare that there is no conflict of interest regarding the publication of this article. AUTHORS’ CONTRIBUTIONS Ton Nu Quynh Trang has conceived of the present idea, carried out andwritten themanuscript with sup- port from Vu Thi Hanh Thu. Ton Nu Quynh Trang and Le Thi Ngoc Tu carried out the experiments in group. TranVanMan has supported the analysis tech- niques. ACKNOWLEDGMENTS This research is funded by the University of Science, VNU-HCM, under grant number T2019-12. REFERENCES 1. 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