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Effect of Cu Addition to 2205 Duplex Stainless Steel on the Resistance against Pitting Corrosion by the Pseudomonas aeruginosa Biofilm
Accepted Manuscript Title: Effect of Cu Addition to 2205 Duplex Stainless Steel on the Resistance Against Pitting Corrosion by the Pseudomonas Aeruginosa Biofilm Author: Ping Li, Yang Zhao, Yuzhi Liu, Ying Zhao, Dake Xu, Chunguang Yang, Tao Zhang, Tingyue Gu, Ke Yang PII: DOI: Reference:
S1005-0302(16)30230-4 http://dx.doi.org/doi: 10.1016/j.jmst.2016.11.020 JMST 855
To appear in:
Journal of Materials Science & Technology
Received date: Revised date: Accepted date:
15-6-2016 8-10-2016 11-11-2016
Please cite this article as: Ping Li, Yang Zhao, Yuzhi Liu, Ying Zhao, Dake Xu, Chunguang Yang, Tao Zhang, Tingyue Gu, Ke Yang, Effect of Cu Addition to 2205 Duplex Stainless Steel on the Resistance Against Pitting Corrosion by the Pseudomonas Aeruginosa Biofilm, Journal of Materials Science & Technology (2016), http://dx.doi.org/doi: 10.1016/j.jmst.2016.11.020. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Effect of Cu Addition to 2205 Duplex Stainless Steel on the Resistance against Pitting Corrosion by the Pseudomonas aeruginosa Biofilm
Ping Li1¶, Yang Zhao1, 2¶, Yuzhi Liu1, Ying Zhao3, Dake Xu2*, Chunguang Yang2, Tao Zhang1, 2*, Tingyue Gu4, Ke Yang2
1
Corrosion and Protection Laboratory, Key Laboratory of Superlight Materials and
Surface Technology, Harbin Engineering University, Ministry of Education, 145 Nantong Street, Harbin 150001, China 2
State Key Laboratory for Corrosion and Protection, Institute of Metal Research,
Chinese Academy of Sciences, Shenyang 110016, China 3
Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences,
Shenzhen, China 4
Department of Chemical and Biomolecular Engineering, Institute for Corrosion and
Multiphase Technology, Ohio University, Athens, Ohio 45701, USA
[Received 15 June 2016; Received in revised form 8 October 2016; Accepted 11 November 2016]
¶ These authors contributed equally to this work. *
Corresponding authors: ([email protected])
Dake
Xu
([email protected]),
Tao
Zhang
¶ These authors contributed equally to this work. 1
Page 1 of 18
Highlights 2205-Cu DSS exhibits better pitting corrosion resistance against P. aeruginosa than 2205 DSS 2205-Cu DSS has a larger CPT value in presence of P. aeruginosa 2205-Cu DSS shows considerably small pits in presence of P. aeruginosa
ABSTRACT
The effect of copper addition to 2205 duplex stainless steel (DSS) on its resistance against pitting corrosion by the Pseudomonas aeruginosa biofilm was investigated using electrochemical and surface analysis techniques. Cu addition decreased the general corrosion resistance, resulting in a higher general corrosion rate in the sterile medium. Because DSS usually has a very small general corrosion rate, its pitting corrosion resistance is far more important. In this work, it was shown that 2205-3%Cu DSS exhibited a much higher pitting corrosion resistance against the P. aeruginosa biofilm compared with the 2205 DSS control, characterized by no significant change in the pitting potential and critical pitting temperature (CPT) values. The strong pitting resistance ability of 2205-3%Cu DSS could be attributed to the copper-rich phases on the surface and the release of copper ions, providing a strong antibacterial ability that inhibited the attachment and growth of the corrosive P. aeruginosa biofilm. Keywords: Duplex stainless steel; Copper; Microbiologically influenced corrosion; Pitting corrosion; Antibacterial; Pseudomonas aeruginosa 2
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1. Introduction Duplex stainless steels (DSSs) are highly resistant against conventional corrosion. DSSs contain both ferrite and austenite, and large amounts of key passivating elements (Cr, Mo and Ni) [1]. Thus, DSSs have wide applications in the marine and petrochemical industries [2]. However, DSSs suffer from severe pitting corrosion caused by microorganisms, known as microbiologically influenced corrosion (MIC). Antony et al. [3] found that sulphate-reducing bacteria (SRB) would modify the passive film locally leading to pitting corrosion. Moradi et al. [4] found that a high concentration of chloride ion was present in the biofilm structure, causing a loss of chromium compounds underneath the biofilm and thus resulting in MIC pitting corrosion.
Pseudomonas aeruginosa is an aerobic, Gram-negative motile rod-shaped bacterium, which has been frequently found in gasoline tanks and marine environments [5]. Many researchers confirmed that P. aeruginosa was capable of inducing the pitting corrosion of carbon steels and stainless steels (SSs) [6-7]. It was 3
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reported that copper addition inhibited the pitting corrosion of 304 SS against the Escherichia coli biofilm [8].
Just like the ductile-brittle transition temperature, there is a critical temperature for the pitting corrosion, which is one of the factors used to rank the pitting corrosion resistance of SSs. The concept of critical pitting temperature (CPT) was introduced by Brigham and Tozer [9]. Below the CPT, stable pitting does not take place. Once the temperature exceeds CPT, the passive film breaks down and stable pitting occurs. So far, there is no reported information on the effects of Cu addition to DSSs on the pitting corrosion resistance of the alloy against biofilms. For example, it remains to see whether bacteria influence the CPT. It was reported that Cu decreased the abiotic pitting corrosion resistance of DSSs [10-11], but can the Cu addition also decrease the CPT of DSSs in the presence of bacteria? This work was the first attempt to answer these questions.
2. Experimental 2.1. Metal Materials The 2205-3%Cu DSS was melted using a vacuum furnace, casted into ingots, and then hot-rolled into 10 mm thick plates. The metal specimens were first annealed at 1050 °C for 1 h and then quenched in water. Finally, the specimens were aged at 540 °C for 4 h to precipitate the Cu-rich phase. The commercial 2205 DSS (Taiyuan Iron & Steel (Group) Co. Ltd, Taiyuan, Shanxi, China) steel was used for comparison. 4
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The chemical compositions of the DSS steel are given in Table 1. Prior to electrochemical measurements, the specimens were cut into square coupons with dimensions of 10 mm×10 mm×5 mm, then embedded in epoxy resin with an exposed test area of 1 cm2. Each coupon’s surface was wet abraded to a 1000-grit finish, degreased with acetone, rinsed with distilled water and finally dried under a compressed hot air stream.
2.2. Culture medium and inoculum P. aeruginosa (1A00099) was obtained from the Marine Culture Collection of China (MCCC). The 2216E medium used in this study contained 19.45 g/L NaCl, 5.98 g/L MgCl2, 3.24 g/L Na2SO4, 1.8 g/L CaCl2, 0.55 g/L KCl, 0.16 g/L Na2CO3, 0.08 g/L KBr, 0.034 g/L SrCl2, 0.08 g/L SrBr2, 0.022 g/L H3BO, 0.004 g/L NaSiO3, 0.0024 g/L NaF, 0.0016 g/L NH4NO3, 0.008 g/L Na2HPO4, 5.0 g/L peptone, 1.0 g/L yeast extract and 0.1 g/L ferric citrate. The initial cell concentration was approximately 106 cells mL-1 right after inoculation and the pH of the medium was adjusted to 7.6 ± 0.2.
2.3. Electrochemical measurements For the measurements of polarization curves, a three-electrode cell setup was used with a standard calomel electrode (SCE) as the reference electrode and a platinum counter electrode (20 mm × 20 mm). Potentiodynamic polarization tests were carried out in the 2216E culture medium. The scan rate was 0.166 mV s-1 and 5
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the starting potential was −0.3 V vs. the open-circuit potential. The test was terminated when the current exceeded 1 mA. Electrochemical noise (EN) measurements were carried out with a Zahner IM6e impedance measurement unit equipped with a Zahner NProbe (IM6, Germany). Two identical specimens were used as the working electrode and an Ag/AgCl electrode as the reference electrode, respectively. EN data were instantaneously recorded for 4800 s at a collection frequency of 4 Hz. The medium temperature during the test increased from 10 to 80 o
C, and the scanning rate was 1 oC min-1 controlled by a programmable temperature
controller (THCD-09, Zjnbth, China). The direct current trend of the potential and current noise data were removed using the 5th-order polynomial detrending method [12]. CPT values in each experiment were determined using a new criterion method reported by Zhang et al. [13]: the calculation was based on the 1/Rn values and the replotted Arrhenius plot. Once the temperature exceeded the transition point, the activation energy of corrosion changed to a negative value, and the corrosion events became spontaneous processes. This transition point corresponded to CPT.
2.4. Microscopic observation To observe the surface morphology of 2205 DSS and 2205-3%Cu DSS, the specimens were abraded successively to 2000-grit finish, polished with a 0.25 μm diamond powder paste, and then electrochemically etched in a 30 wt% KOH solution, which revealed the microstructures with austenite phase and ferrite phase. The volume fractions of ferrite and austenite were evaluated using an X-ray diffraction (XRD) 6
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machine (SHIMADZU XRD-7000 S/L, Japan) with a CuKα X-ray source operated at 40 kV and 150 mA. Scans were performed over a range 20°--80° using a step size of 0.333 s.
The biofilm morphologies of the 2205 DSS specimens after incubation for 14 days in P. aeruginosa inoculated medium were observed using confocal laser scanning microscopy (CLSM, Model C2si+; Nikon Co., Kyoto, Japan). All coupons were stained with 4', 6-diamidino-2-phenylindole (DAPI, Sigma-Aldrich Co., MO, USA) to evaluate the antibacterial performance of 2205-3%Cu DSS [14]. The maximum depth of the pits were measured under a Zeiss confocal laser scanning microscope (CLSM) (LSM 710, Zeiss, Jena, Germany) after incubation for 14 days.
2.5 Measurement of copper ion release The coupons were immersed in 5 mL centrifuge tubes containing 2 mL sterilized 2216E medium. The Cu2+ concentration was measured after incubation for 1, 3, 7, 14 days, respectively, by using atomic absorption spectroscopy (AAS) (Z-2000, Hitachi, Tokyo, Japan). A graphite furnace was adopted for atomization. Air-C2H2 was used as the carrier gas at a flow rate of 1.8l min−1. The flow rate of the oxidant gas was 15.0 l min−1 at a pressure of 160 kPa. The lamp current was 280 mA, the wave length and the slit width were 324.8 nm and 1.3 nm, respectively.
3. Results 7
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3.1 Microstructure Representative micrographs of 2205 DSS and 2205-3%Cu DSS cross-section surfaces are presented in Fig. 1(a) and (b). It can be seen that the microstructure of 2205 DSS and 2205-3%Cu DSS consisted of austenite (bright) and ferrite (dark) phases, each elongating in the rolling direction. The interface between the two-phases was clearly etched, and the island-like austenite phase was embedded in the continuous ferrite matrix. The volume fraction of austenite in 2205-3%Cu DSS matrix was increased up to 60% according to the XRD spectra calculation as shown in Fig. 2, which might be caused by the addition of copper, a well known austenite stabilizer, increasing the austenite fraction in the intercritical zone and enlarging the area of the austenite during the metallurgical process [15].
3.2 Potentiodynamic polarization and CPT measurement results Fig. 3(a) shows the potentiodynamic polarization curves for 2205 DSS and 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium. The electrochemical parameters from fitting with Tafel extrapolation, are shown in Table 2. When incubated with P. aeruginosa, the pitting potential of 2205 DSS decreased by 393 mV. Furthermore, both the anodic and the cathodic reactions were accelerated and the corrosion current and passive current of 2205 DSS increased to 1.28 μA cm−2 and 2.86 μA cm−2, respectively, which were nearly 4 times higher than those of 2205 DSS in the sterile medium. This result implied that the P. aeruginosa biofilm probably destroyed the passive film and decreased the pitting resistance of 8
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2205 DSS [16]. With the addition of Cu to 2205 DSS, the polarization curves of 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium shifted to the right compared with those of 2205 DSS, indicating the increase of corrosion current and passive current, which could be attributed to the increase of the volume fraction of the austenite introduced by the copper element. However, the pitting potential of 2205-3%Cu DSS in the P. aeruginosa inoculated medium only decreased slightly (about 30 mV) compared with that of 2205 DSS, suggesting that the presence of Cu improved the pitting corrosion resistance of DSS.
Fig. 3(b) depicts the development of CPT of 2205 DSS and 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium after various immersion times. In the case of the sterile medium, with the addition of Cu element, the CPT of 2205-3%Cu DSS decreased from approximately 65 to 55 °C, suggesting that the presence of Cu element increased the pitting susceptibility of 2205-3%Cu DSS. When incubated with P. aeruginosa, however, the CPT of the 2205 DSS was 60 °C after the 1-day immersion, and then decreased to 45 °C after 14 days of immersion. In comparison, the CPT of 2205-3%Cu DSS was almost not affected by P. aeruginosa, only decreasing from 57 to 54 °C in the first day, and then remained at 54 °C (much larger than 45 °C) for the whole immersion period of 14 days. These results suggested that the addition of Cu increased the pitting corrosion resistance of 2205-3%Cu DSS in the P. aeruginosa inoculated medium.
9
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3.3 Corrosion morphology observation The P. aeruginosa biofilms on the 2205 DSS and 2205-3%Cu DSS coupons were shown in Fig. 4. It can be seen that the biofilm on the 2205-3%Cu DSS surface was much less than that on the 2205 DSS surface after 14-day immersion. This could be attributed to the copper’s antibacterial ability that inhibited the P. aeruginosa biofilm.
After the biofilms were removed, the maximum depth of pits obtained from 3-D CLSM images for 2205 DSS and 2205-3%Cu DSS coupons in the P. aeruginosa inoculated medium after 14 days. The images are shown in Fig. 5. The largest pit depth for 2205 DSS was 4.83 μm with a surface diameter of 10.84 μm as shown in Fig. 5(a). In comparison，the largest pit depth for 2205-3%Cu DSS was only 1.53 μm with a surface diameter of 8.61 μm as shown in Fig. 5(b). Fig. 5 confirms that the addition of Cu element to 2205 DSS increased its pitting corrosion resistance against P. aeruginosa, which was consistent with the CPT results above.
3.4 Concentration of the released copper ion Fig. 6 shows the concentration of the release of Cu2+ from 2205-3%Cu DSS into the sterile medium for different immersion times. On the first day, the Cu2+ concentration reached 17.6 ppb and it decreased sharply to 14.3 ppb after three days. After three days, the Cu2+ concentration started to increase, reaching 19.4 ppb after 14 days. This indicates that the 2205 Cu-DSS matrix released Cu2+ into the medium and maintained a relative higher Cu2+ concentration in the medium. 10
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4. Discussion 4.1 Influence of Cu element addition to 2205 DSS on its corrosion resistance The antibacterial effect of Cu2+ enhanced the MIC pitting resistance. However, addition of Cu to 2205 DSS added an adverse effect on the pitting resistance equivalent number (PREN). PREN is often used to rank the pitting resistance of DSS, which is strongly dependent on the contents of three most important elements: chromium, molybdenum and nitrogen [17]. Copper is an element that expands the volume fraction of austenite. Thus, 2205-3%Cu DSS possesses a higher percentage of austenite, and a lower PREN value of austenite (36.2) compared with that of ferrite (36.7) when annealed at the temperature of 1050 °C [18]. This suggests that the austenite phases are the weaker phase where stable pitting may occur preferentially. Thus, the Cu element addition to 2205 DSS increased its general corrosion rate both in the sterile medium and in the P. aeruginosa inoculated medium as shown in Table 1 and Fig. 3(a).
4.2 Influence of Cu element addition to 2205 DSS on its CPT Zahner NProbe Because MIC pitting is caused by corrosive biofilms, successful biofilm inhibition may result in effective MIC pitting mitigation. When the concentration of the release of Cu2+ ions increased, the biofilm was gradually inhibited over time. The 2205-3%Cu DSS induced the death of the sessile P. aeruginosa cells, especially those that were directly attached to the metal surface after 11
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14 days of incubation [19] as shown in Fig. 4. The dead biofilm decreased the probability of the occurrence of stable pitting [19,20], and thus the maximum MIC pit depth on 2205-3%Cu DSS coupons were all significantly reduced (Fig. 5), suggesting that Cu addition to 2205 DSS increased its MIC pitting resistance.
5. Conclusions Copper had an important influence on the pitting corrosion resistance of the 2205-3%Cu DSS against the P. aeruginosa biofilm. 2205-3%Cu DSS exhibited higher MIC pitting resistance, resulting in considerably smaller pit depth and no significant change in pitting potential and CPT values. This was because Cu2+ ions released from 2205-3%Cu DSS possessed a strong antibacterial ability that inhibited the corrosive P. aeruginosa biofilm.
On the other hand, copper increased the 2205-3%Cu DSS general corrosion rate both in the sterile medium and in the P. aeruginosa inoculated medium, because copper expanded the area of austenite that had a lower PREN value. Because MIC pitting resistance of DSS is more important than the general corrosion rate which is usually very small, the addition of Cu allows 2205-3%Cu DSS for use in environments that MIC pitting presents a real threat.
Acknowledgements The authors wish to acknowledge the financial support of the program of Outstanding 12
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Young Scholars, the National Natural Science Foundation of China (No. 51371182). This work was financially supported by Shenzhen Science and Technology Research funding (JCYJ20160608153641020), the National Basic Research Program of China (973 Program Project No. 2014CB643300), the National Natural Science Foundation (No. 51501203 and U1660118), the National Environmental Corrosion Platform (NECP) and the “Young Merit Scholars” program of the Institute of Metal Research, Chinese Academy of Sciences.
References [1] L.L. Machuca, S.I. Bailey, R. Gubner, E.L.J. Watkin, M.P. Ginige, A.H. Kaksonen, K. Heidersbach, Corros. Sci. 67 (2013) 242-255. [2] P.J. Antony, R.K.S. Raman, P. Kumar, R. Raman, Corros. Sci. 52 (2010) 1404-1412. [3] P.J. Antony, R.K.S. Raman, R. Mohanram, P. Kumar, R. Raman, Corros. Sci. 50 (2008) 1858-1864. [4] M. Moradi, Z.L. Song, L.J. Yang, J.J. Jiang, J. He, Corros. Sci. 84 (2014) 103-112. [5] H.B. Li, E.Z. Zhou, Y.B. Ren, D.W. Zhang, D.K. Xu, C.G. Yang, H. Feng, Z.H. Jiang, X.G. Li, T.Y. Gu, K. Yang, Corros. Sci. 11 (2016) 811-821. [6] D.K. Xu, J. Xia, E.Z. Zhou, D.W. Zhang, H.B. Li, C.G. Yang, Q. Li, H. Lin, X.G. Li, K. Yang, Bioelectrochemistry 22 (2107) 1-8. [7] S.J. Yuan, S.O. Pehkonen, Corros. Sci. 51 (2009) 1372-1385. [8] D. Sun, M.B. Shahzad, M. Li, G. Wang, D. Xu, Mater. Technol. 30 (2015) 13
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b90-b95. [9] R. Brigham, E. Tozer, Corrosion 30 (1974) 161-166. [10] Q. Ran, J. Li, Y. Xu, X. Xiao, H. Yu, L. Jiang, Mater. Des. 46 (2013) 758-765. [11] T. Ujiro, S. Satoh, R.W. Staehle, W.H. Smyrl, Corros. Sci. 43 (2001) 185-200. [12] R.A. Cottis, Corrosion 57 (2001) 265-276. [13] T. Zhang, D.Y. Wang, Y.W. Shao, G.Z. Meng, F.H. Wang, Corros. Sci. 58 (2012) 202–210. [14] D. Sun D.K. Xu C.G. Yang, J. Chen, M. Babar Shahzad, Z.Q. Sun, J.L. Zhao, T.Y. Gu, K. Yang, G.X. Wang, Mater. Sci. Eng. C 69 (2016) 744-750. [15] L. Nan, G.A. Ren, D.H. Wang, K. Yang, J. Mater. Sci. Technol. 5 (2016) 445-451. [16] H.B. Li, E.Z. Zhou, D.W. Zhang, D.K. Xu, J. Xia, C.G. Yang, H. Feng, Z.H. Jiang, X.G.
Li, T.Y. Gu, K. Yang, Scientific Reports 2 (2015) 1-12.
[17] M.H. Moayed, R.C. Newman, Corros. Sci. 48 (2016) 3513-3530. [18] Z.Y. Zhang, Z.Y. Wang, Y.M. Jiang, H. Tan, D. Han, Y.J. Guo, J. Li, Corros. Sci. 62 (2012) 42-50. [19] J. Xia, C.G. Yang, D.K. Xu, D. Sun, L. Nan, Z.Q. Sun, Q. Li, T.Y. Gu, K. Yang, Biofouling 31 (2015) 481-491. [20] L. Nan, Y. Liu, M. Lü, K. Yang, J. Mater. Sci.-Mater. M 19 (2008) 3057-3062.
14
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List of Figures： Fig. 1. Optical images of microstructures of: (a) 2205 DSS and (b) 2205-3%Cu DSS. Fig. 2. XRD spectra of 2205 and 2205-3%Cu DSS. Fig. 3. Electrochemical characteristics of 2205 DSS and 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium: (a) potentiodynamic polarization curves for 1 day, (b) CPT values for 14 days of immersion. Fig. 4. CLSM images of DAPI stained biofilms on: (a) 2205 and (b) 2205-3%Cu DSS coupons after 14-day immersion in P. aeruginosa inoculated medium. Blue dots represent the bacteria cells. Fig. 5. Largest pit depth measured under CLSM of: (a) 2205 and (b) 2205-3%Cu DSS in the P. aeruginosa inoculated medium after 14 days. Fig. 6. Time course of Cu2+ release from 2205-3%Cu DSS in the sterile medium.
15
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Table 1. Chemical compositions of 2205 DSS and 2205-3%Cu DSS. Component
Si
Mn
P
S
Ni
Cr
Mo
DSS (wt%) 0.51 1.14 0.030 <0.0010 5.89 23.22 3.10
Cu -
N
Fe
C
0.17 Bal. <0.03
Cu-DSS 0.04 0.01 0.006
0.0034
6.03 23.63 2.90 3.02 0.23 Bal. <0.03
(wt%)
Table 2. Fitted electrochemical parameters for 2205 DSS and 2205-3%Cu DSS in the sterile and the P. aeruginosa inoculated mediums. Ecorr (mV) 2205 in sterile medium 2205 in the P. aeruginosa inoculated medium 2205-3%Cu in sterile medium 2205-3%Cu in the P. aeruginosa inoculated medium
icorr (μA/cm2)
ip (μA/cm2)
Epit (mV)
-50
0.39
0.74
1505
19.6
1.28
2.86
1112
220
1.49
6.25
1405
160
3.12
13.9
1372
16
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Fig. 1
Fig. 2
Fig. 3
17
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Fig. 4
Distance (μm)
Distance (μm)
Fig. 5
Fig. 6
18
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S1005-0302(16)30230-4 http://dx.doi.org/doi: 10.1016/j.jmst.2016.11.020 JMST 855
To appear in:
Journal of Materials Science & Technology
Received date: Revised date: Accepted date:
15-6-2016 8-10-2016 11-11-2016
Please cite this article as: Ping Li, Yang Zhao, Yuzhi Liu, Ying Zhao, Dake Xu, Chunguang Yang, Tao Zhang, Tingyue Gu, Ke Yang, Effect of Cu Addition to 2205 Duplex Stainless Steel on the Resistance Against Pitting Corrosion by the Pseudomonas Aeruginosa Biofilm, Journal of Materials Science & Technology (2016), http://dx.doi.org/doi: 10.1016/j.jmst.2016.11.020. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Effect of Cu Addition to 2205 Duplex Stainless Steel on the Resistance against Pitting Corrosion by the Pseudomonas aeruginosa Biofilm
Ping Li1¶, Yang Zhao1, 2¶, Yuzhi Liu1, Ying Zhao3, Dake Xu2*, Chunguang Yang2, Tao Zhang1, 2*, Tingyue Gu4, Ke Yang2
1
Corrosion and Protection Laboratory, Key Laboratory of Superlight Materials and
Surface Technology, Harbin Engineering University, Ministry of Education, 145 Nantong Street, Harbin 150001, China 2
State Key Laboratory for Corrosion and Protection, Institute of Metal Research,
Chinese Academy of Sciences, Shenyang 110016, China 3
Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences,
Shenzhen, China 4
Department of Chemical and Biomolecular Engineering, Institute for Corrosion and
Multiphase Technology, Ohio University, Athens, Ohio 45701, USA
[Received 15 June 2016; Received in revised form 8 October 2016; Accepted 11 November 2016]
¶ These authors contributed equally to this work. *
Corresponding authors: ([email protected])
Dake
Xu
([email protected]),
Tao
Zhang
¶ These authors contributed equally to this work. 1
Page 1 of 18
Highlights 2205-Cu DSS exhibits better pitting corrosion resistance against P. aeruginosa than 2205 DSS 2205-Cu DSS has a larger CPT value in presence of P. aeruginosa 2205-Cu DSS shows considerably small pits in presence of P. aeruginosa
ABSTRACT
The effect of copper addition to 2205 duplex stainless steel (DSS) on its resistance against pitting corrosion by the Pseudomonas aeruginosa biofilm was investigated using electrochemical and surface analysis techniques. Cu addition decreased the general corrosion resistance, resulting in a higher general corrosion rate in the sterile medium. Because DSS usually has a very small general corrosion rate, its pitting corrosion resistance is far more important. In this work, it was shown that 2205-3%Cu DSS exhibited a much higher pitting corrosion resistance against the P. aeruginosa biofilm compared with the 2205 DSS control, characterized by no significant change in the pitting potential and critical pitting temperature (CPT) values. The strong pitting resistance ability of 2205-3%Cu DSS could be attributed to the copper-rich phases on the surface and the release of copper ions, providing a strong antibacterial ability that inhibited the attachment and growth of the corrosive P. aeruginosa biofilm. Keywords: Duplex stainless steel; Copper; Microbiologically influenced corrosion; Pitting corrosion; Antibacterial; Pseudomonas aeruginosa 2
Page 2 of 18
1. Introduction Duplex stainless steels (DSSs) are highly resistant against conventional corrosion. DSSs contain both ferrite and austenite, and large amounts of key passivating elements (Cr, Mo and Ni) [1]. Thus, DSSs have wide applications in the marine and petrochemical industries [2]. However, DSSs suffer from severe pitting corrosion caused by microorganisms, known as microbiologically influenced corrosion (MIC). Antony et al. [3] found that sulphate-reducing bacteria (SRB) would modify the passive film locally leading to pitting corrosion. Moradi et al. [4] found that a high concentration of chloride ion was present in the biofilm structure, causing a loss of chromium compounds underneath the biofilm and thus resulting in MIC pitting corrosion.
Pseudomonas aeruginosa is an aerobic, Gram-negative motile rod-shaped bacterium, which has been frequently found in gasoline tanks and marine environments [5]. Many researchers confirmed that P. aeruginosa was capable of inducing the pitting corrosion of carbon steels and stainless steels (SSs) [6-7]. It was 3
Page 3 of 18
reported that copper addition inhibited the pitting corrosion of 304 SS against the Escherichia coli biofilm [8].
Just like the ductile-brittle transition temperature, there is a critical temperature for the pitting corrosion, which is one of the factors used to rank the pitting corrosion resistance of SSs. The concept of critical pitting temperature (CPT) was introduced by Brigham and Tozer [9]. Below the CPT, stable pitting does not take place. Once the temperature exceeds CPT, the passive film breaks down and stable pitting occurs. So far, there is no reported information on the effects of Cu addition to DSSs on the pitting corrosion resistance of the alloy against biofilms. For example, it remains to see whether bacteria influence the CPT. It was reported that Cu decreased the abiotic pitting corrosion resistance of DSSs [10-11], but can the Cu addition also decrease the CPT of DSSs in the presence of bacteria? This work was the first attempt to answer these questions.
2. Experimental 2.1. Metal Materials The 2205-3%Cu DSS was melted using a vacuum furnace, casted into ingots, and then hot-rolled into 10 mm thick plates. The metal specimens were first annealed at 1050 °C for 1 h and then quenched in water. Finally, the specimens were aged at 540 °C for 4 h to precipitate the Cu-rich phase. The commercial 2205 DSS (Taiyuan Iron & Steel (Group) Co. Ltd, Taiyuan, Shanxi, China) steel was used for comparison. 4
Page 4 of 18
The chemical compositions of the DSS steel are given in Table 1. Prior to electrochemical measurements, the specimens were cut into square coupons with dimensions of 10 mm×10 mm×5 mm, then embedded in epoxy resin with an exposed test area of 1 cm2. Each coupon’s surface was wet abraded to a 1000-grit finish, degreased with acetone, rinsed with distilled water and finally dried under a compressed hot air stream.
2.2. Culture medium and inoculum P. aeruginosa (1A00099) was obtained from the Marine Culture Collection of China (MCCC). The 2216E medium used in this study contained 19.45 g/L NaCl, 5.98 g/L MgCl2, 3.24 g/L Na2SO4, 1.8 g/L CaCl2, 0.55 g/L KCl, 0.16 g/L Na2CO3, 0.08 g/L KBr, 0.034 g/L SrCl2, 0.08 g/L SrBr2, 0.022 g/L H3BO, 0.004 g/L NaSiO3, 0.0024 g/L NaF, 0.0016 g/L NH4NO3, 0.008 g/L Na2HPO4, 5.0 g/L peptone, 1.0 g/L yeast extract and 0.1 g/L ferric citrate. The initial cell concentration was approximately 106 cells mL-1 right after inoculation and the pH of the medium was adjusted to 7.6 ± 0.2.
2.3. Electrochemical measurements For the measurements of polarization curves, a three-electrode cell setup was used with a standard calomel electrode (SCE) as the reference electrode and a platinum counter electrode (20 mm × 20 mm). Potentiodynamic polarization tests were carried out in the 2216E culture medium. The scan rate was 0.166 mV s-1 and 5
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the starting potential was −0.3 V vs. the open-circuit potential. The test was terminated when the current exceeded 1 mA. Electrochemical noise (EN) measurements were carried out with a Zahner IM6e impedance measurement unit equipped with a Zahner NProbe (IM6, Germany). Two identical specimens were used as the working electrode and an Ag/AgCl electrode as the reference electrode, respectively. EN data were instantaneously recorded for 4800 s at a collection frequency of 4 Hz. The medium temperature during the test increased from 10 to 80 o
C, and the scanning rate was 1 oC min-1 controlled by a programmable temperature
controller (THCD-09, Zjnbth, China). The direct current trend of the potential and current noise data were removed using the 5th-order polynomial detrending method [12]. CPT values in each experiment were determined using a new criterion method reported by Zhang et al. [13]: the calculation was based on the 1/Rn values and the replotted Arrhenius plot. Once the temperature exceeded the transition point, the activation energy of corrosion changed to a negative value, and the corrosion events became spontaneous processes. This transition point corresponded to CPT.
2.4. Microscopic observation To observe the surface morphology of 2205 DSS and 2205-3%Cu DSS, the specimens were abraded successively to 2000-grit finish, polished with a 0.25 μm diamond powder paste, and then electrochemically etched in a 30 wt% KOH solution, which revealed the microstructures with austenite phase and ferrite phase. The volume fractions of ferrite and austenite were evaluated using an X-ray diffraction (XRD) 6
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machine (SHIMADZU XRD-7000 S/L, Japan) with a CuKα X-ray source operated at 40 kV and 150 mA. Scans were performed over a range 20°--80° using a step size of 0.333 s.
The biofilm morphologies of the 2205 DSS specimens after incubation for 14 days in P. aeruginosa inoculated medium were observed using confocal laser scanning microscopy (CLSM, Model C2si+; Nikon Co., Kyoto, Japan). All coupons were stained with 4', 6-diamidino-2-phenylindole (DAPI, Sigma-Aldrich Co., MO, USA) to evaluate the antibacterial performance of 2205-3%Cu DSS [14]. The maximum depth of the pits were measured under a Zeiss confocal laser scanning microscope (CLSM) (LSM 710, Zeiss, Jena, Germany) after incubation for 14 days.
2.5 Measurement of copper ion release The coupons were immersed in 5 mL centrifuge tubes containing 2 mL sterilized 2216E medium. The Cu2+ concentration was measured after incubation for 1, 3, 7, 14 days, respectively, by using atomic absorption spectroscopy (AAS) (Z-2000, Hitachi, Tokyo, Japan). A graphite furnace was adopted for atomization. Air-C2H2 was used as the carrier gas at a flow rate of 1.8l min−1. The flow rate of the oxidant gas was 15.0 l min−1 at a pressure of 160 kPa. The lamp current was 280 mA, the wave length and the slit width were 324.8 nm and 1.3 nm, respectively.
3. Results 7
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3.1 Microstructure Representative micrographs of 2205 DSS and 2205-3%Cu DSS cross-section surfaces are presented in Fig. 1(a) and (b). It can be seen that the microstructure of 2205 DSS and 2205-3%Cu DSS consisted of austenite (bright) and ferrite (dark) phases, each elongating in the rolling direction. The interface between the two-phases was clearly etched, and the island-like austenite phase was embedded in the continuous ferrite matrix. The volume fraction of austenite in 2205-3%Cu DSS matrix was increased up to 60% according to the XRD spectra calculation as shown in Fig. 2, which might be caused by the addition of copper, a well known austenite stabilizer, increasing the austenite fraction in the intercritical zone and enlarging the area of the austenite during the metallurgical process [15].
3.2 Potentiodynamic polarization and CPT measurement results Fig. 3(a) shows the potentiodynamic polarization curves for 2205 DSS and 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium. The electrochemical parameters from fitting with Tafel extrapolation, are shown in Table 2. When incubated with P. aeruginosa, the pitting potential of 2205 DSS decreased by 393 mV. Furthermore, both the anodic and the cathodic reactions were accelerated and the corrosion current and passive current of 2205 DSS increased to 1.28 μA cm−2 and 2.86 μA cm−2, respectively, which were nearly 4 times higher than those of 2205 DSS in the sterile medium. This result implied that the P. aeruginosa biofilm probably destroyed the passive film and decreased the pitting resistance of 8
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2205 DSS [16]. With the addition of Cu to 2205 DSS, the polarization curves of 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium shifted to the right compared with those of 2205 DSS, indicating the increase of corrosion current and passive current, which could be attributed to the increase of the volume fraction of the austenite introduced by the copper element. However, the pitting potential of 2205-3%Cu DSS in the P. aeruginosa inoculated medium only decreased slightly (about 30 mV) compared with that of 2205 DSS, suggesting that the presence of Cu improved the pitting corrosion resistance of DSS.
Fig. 3(b) depicts the development of CPT of 2205 DSS and 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium after various immersion times. In the case of the sterile medium, with the addition of Cu element, the CPT of 2205-3%Cu DSS decreased from approximately 65 to 55 °C, suggesting that the presence of Cu element increased the pitting susceptibility of 2205-3%Cu DSS. When incubated with P. aeruginosa, however, the CPT of the 2205 DSS was 60 °C after the 1-day immersion, and then decreased to 45 °C after 14 days of immersion. In comparison, the CPT of 2205-3%Cu DSS was almost not affected by P. aeruginosa, only decreasing from 57 to 54 °C in the first day, and then remained at 54 °C (much larger than 45 °C) for the whole immersion period of 14 days. These results suggested that the addition of Cu increased the pitting corrosion resistance of 2205-3%Cu DSS in the P. aeruginosa inoculated medium.
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3.3 Corrosion morphology observation The P. aeruginosa biofilms on the 2205 DSS and 2205-3%Cu DSS coupons were shown in Fig. 4. It can be seen that the biofilm on the 2205-3%Cu DSS surface was much less than that on the 2205 DSS surface after 14-day immersion. This could be attributed to the copper’s antibacterial ability that inhibited the P. aeruginosa biofilm.
After the biofilms were removed, the maximum depth of pits obtained from 3-D CLSM images for 2205 DSS and 2205-3%Cu DSS coupons in the P. aeruginosa inoculated medium after 14 days. The images are shown in Fig. 5. The largest pit depth for 2205 DSS was 4.83 μm with a surface diameter of 10.84 μm as shown in Fig. 5(a). In comparison，the largest pit depth for 2205-3%Cu DSS was only 1.53 μm with a surface diameter of 8.61 μm as shown in Fig. 5(b). Fig. 5 confirms that the addition of Cu element to 2205 DSS increased its pitting corrosion resistance against P. aeruginosa, which was consistent with the CPT results above.
3.4 Concentration of the released copper ion Fig. 6 shows the concentration of the release of Cu2+ from 2205-3%Cu DSS into the sterile medium for different immersion times. On the first day, the Cu2+ concentration reached 17.6 ppb and it decreased sharply to 14.3 ppb after three days. After three days, the Cu2+ concentration started to increase, reaching 19.4 ppb after 14 days. This indicates that the 2205 Cu-DSS matrix released Cu2+ into the medium and maintained a relative higher Cu2+ concentration in the medium. 10
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4. Discussion 4.1 Influence of Cu element addition to 2205 DSS on its corrosion resistance The antibacterial effect of Cu2+ enhanced the MIC pitting resistance. However, addition of Cu to 2205 DSS added an adverse effect on the pitting resistance equivalent number (PREN). PREN is often used to rank the pitting resistance of DSS, which is strongly dependent on the contents of three most important elements: chromium, molybdenum and nitrogen [17]. Copper is an element that expands the volume fraction of austenite. Thus, 2205-3%Cu DSS possesses a higher percentage of austenite, and a lower PREN value of austenite (36.2) compared with that of ferrite (36.7) when annealed at the temperature of 1050 °C [18]. This suggests that the austenite phases are the weaker phase where stable pitting may occur preferentially. Thus, the Cu element addition to 2205 DSS increased its general corrosion rate both in the sterile medium and in the P. aeruginosa inoculated medium as shown in Table 1 and Fig. 3(a).
4.2 Influence of Cu element addition to 2205 DSS on its CPT Zahner NProbe Because MIC pitting is caused by corrosive biofilms, successful biofilm inhibition may result in effective MIC pitting mitigation. When the concentration of the release of Cu2+ ions increased, the biofilm was gradually inhibited over time. The 2205-3%Cu DSS induced the death of the sessile P. aeruginosa cells, especially those that were directly attached to the metal surface after 11
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14 days of incubation [19] as shown in Fig. 4. The dead biofilm decreased the probability of the occurrence of stable pitting [19,20], and thus the maximum MIC pit depth on 2205-3%Cu DSS coupons were all significantly reduced (Fig. 5), suggesting that Cu addition to 2205 DSS increased its MIC pitting resistance.
5. Conclusions Copper had an important influence on the pitting corrosion resistance of the 2205-3%Cu DSS against the P. aeruginosa biofilm. 2205-3%Cu DSS exhibited higher MIC pitting resistance, resulting in considerably smaller pit depth and no significant change in pitting potential and CPT values. This was because Cu2+ ions released from 2205-3%Cu DSS possessed a strong antibacterial ability that inhibited the corrosive P. aeruginosa biofilm.
On the other hand, copper increased the 2205-3%Cu DSS general corrosion rate both in the sterile medium and in the P. aeruginosa inoculated medium, because copper expanded the area of austenite that had a lower PREN value. Because MIC pitting resistance of DSS is more important than the general corrosion rate which is usually very small, the addition of Cu allows 2205-3%Cu DSS for use in environments that MIC pitting presents a real threat.
Acknowledgements The authors wish to acknowledge the financial support of the program of Outstanding 12
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Young Scholars, the National Natural Science Foundation of China (No. 51371182). This work was financially supported by Shenzhen Science and Technology Research funding (JCYJ20160608153641020), the National Basic Research Program of China (973 Program Project No. 2014CB643300), the National Natural Science Foundation (No. 51501203 and U1660118), the National Environmental Corrosion Platform (NECP) and the “Young Merit Scholars” program of the Institute of Metal Research, Chinese Academy of Sciences.
References [1] L.L. Machuca, S.I. Bailey, R. Gubner, E.L.J. Watkin, M.P. Ginige, A.H. Kaksonen, K. Heidersbach, Corros. Sci. 67 (2013) 242-255. [2] P.J. Antony, R.K.S. Raman, P. Kumar, R. Raman, Corros. Sci. 52 (2010) 1404-1412. [3] P.J. Antony, R.K.S. Raman, R. Mohanram, P. Kumar, R. Raman, Corros. Sci. 50 (2008) 1858-1864. [4] M. Moradi, Z.L. Song, L.J. Yang, J.J. Jiang, J. He, Corros. Sci. 84 (2014) 103-112. [5] H.B. Li, E.Z. Zhou, Y.B. Ren, D.W. Zhang, D.K. Xu, C.G. Yang, H. Feng, Z.H. Jiang, X.G. Li, T.Y. Gu, K. Yang, Corros. Sci. 11 (2016) 811-821. [6] D.K. Xu, J. Xia, E.Z. Zhou, D.W. Zhang, H.B. Li, C.G. Yang, Q. Li, H. Lin, X.G. Li, K. Yang, Bioelectrochemistry 22 (2107) 1-8. [7] S.J. Yuan, S.O. Pehkonen, Corros. Sci. 51 (2009) 1372-1385. [8] D. Sun, M.B. Shahzad, M. Li, G. Wang, D. Xu, Mater. Technol. 30 (2015) 13
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b90-b95. [9] R. Brigham, E. Tozer, Corrosion 30 (1974) 161-166. [10] Q. Ran, J. Li, Y. Xu, X. Xiao, H. Yu, L. Jiang, Mater. Des. 46 (2013) 758-765. [11] T. Ujiro, S. Satoh, R.W. Staehle, W.H. Smyrl, Corros. Sci. 43 (2001) 185-200. [12] R.A. Cottis, Corrosion 57 (2001) 265-276. [13] T. Zhang, D.Y. Wang, Y.W. Shao, G.Z. Meng, F.H. Wang, Corros. Sci. 58 (2012) 202–210. [14] D. Sun D.K. Xu C.G. Yang, J. Chen, M. Babar Shahzad, Z.Q. Sun, J.L. Zhao, T.Y. Gu, K. Yang, G.X. Wang, Mater. Sci. Eng. C 69 (2016) 744-750. [15] L. Nan, G.A. Ren, D.H. Wang, K. Yang, J. Mater. Sci. Technol. 5 (2016) 445-451. [16] H.B. Li, E.Z. Zhou, D.W. Zhang, D.K. Xu, J. Xia, C.G. Yang, H. Feng, Z.H. Jiang, X.G.
Li, T.Y. Gu, K. Yang, Scientific Reports 2 (2015) 1-12.
[17] M.H. Moayed, R.C. Newman, Corros. Sci. 48 (2016) 3513-3530. [18] Z.Y. Zhang, Z.Y. Wang, Y.M. Jiang, H. Tan, D. Han, Y.J. Guo, J. Li, Corros. Sci. 62 (2012) 42-50. [19] J. Xia, C.G. Yang, D.K. Xu, D. Sun, L. Nan, Z.Q. Sun, Q. Li, T.Y. Gu, K. Yang, Biofouling 31 (2015) 481-491. [20] L. Nan, Y. Liu, M. Lü, K. Yang, J. Mater. Sci.-Mater. M 19 (2008) 3057-3062.
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List of Figures： Fig. 1. Optical images of microstructures of: (a) 2205 DSS and (b) 2205-3%Cu DSS. Fig. 2. XRD spectra of 2205 and 2205-3%Cu DSS. Fig. 3. Electrochemical characteristics of 2205 DSS and 2205-3%Cu DSS in the sterile medium and in the P. aeruginosa inoculated medium: (a) potentiodynamic polarization curves for 1 day, (b) CPT values for 14 days of immersion. Fig. 4. CLSM images of DAPI stained biofilms on: (a) 2205 and (b) 2205-3%Cu DSS coupons after 14-day immersion in P. aeruginosa inoculated medium. Blue dots represent the bacteria cells. Fig. 5. Largest pit depth measured under CLSM of: (a) 2205 and (b) 2205-3%Cu DSS in the P. aeruginosa inoculated medium after 14 days. Fig. 6. Time course of Cu2+ release from 2205-3%Cu DSS in the sterile medium.
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Table 1. Chemical compositions of 2205 DSS and 2205-3%Cu DSS. Component
Si
Mn
P
S
Ni
Cr
Mo
DSS (wt%) 0.51 1.14 0.030 <0.0010 5.89 23.22 3.10
Cu -
N
Fe
C
0.17 Bal. <0.03
Cu-DSS 0.04 0.01 0.006
0.0034
6.03 23.63 2.90 3.02 0.23 Bal. <0.03
(wt%)
Table 2. Fitted electrochemical parameters for 2205 DSS and 2205-3%Cu DSS in the sterile and the P. aeruginosa inoculated mediums. Ecorr (mV) 2205 in sterile medium 2205 in the P. aeruginosa inoculated medium 2205-3%Cu in sterile medium 2205-3%Cu in the P. aeruginosa inoculated medium
icorr (μA/cm2)
ip (μA/cm2)
Epit (mV)
-50
0.39
0.74
1505
19.6
1.28
2.86
1112
220
1.49
6.25
1405
160
3.12
13.9
1372
16
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Fig. 1
Fig. 2
Fig. 3
17
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Fig. 4
Distance (μm)
Distance (μm)
Fig. 5
Fig. 6
18
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