JMEPEG (2015) 24:2688–2698 DOI: 10.1007/s11665-015-1558-2
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Microbially Influenced Corrosion of 304 Stainless Steel and Titanium by P. variotii and A. niger in Humid Atmosphere Dawei Zhang, Feichi Zhou, Kui Xiao, Tianyu Cui, Hongchong Qian, and Xiaogang Li (Submitted December 23, 2014; in revised form April 15, 2015; published online May 21, 2015) Microbially induced corrosion (MIC) poses significant threats to reliability and safety of engineering materials and structures. While most MIC studies focus on prokaryotic bacteria such as sulfate-reducing bacteria, the influence of fungi on corrosion behaviors of metals has not been adequately reported. In this study, 304 stainless steel and titanium were exposed to two very common fungi, Paecilomyces variotii, Aspergillus niger and their mixtures under highly humid atmosphere. The initial corrosion behaviors within 28 days were studied via scanning Kelvin probe, which showed marked surface ennoblement and increasingly heterogeneous potential distribution upon prolonged fungus exposure. Using stereomicroscopy, fungus growth as well as corrosion morphology of 304 stainless steel and titanium were also evaluated after a long-term exposure for 60 days. The presence of fungi decreased the corrosion resistance for both 304 stainless steel and titanium. Titanium showed higher resistance to fungus growth and the induced corrosion. Exposure to the mixed strains resulted in the highest fungus growth rate but the mildest corrosion, possibly due to the decreased oxygen level by increased fungal activities. Keywords
advanced characterization, atmospheric corrosion, corrosion and wear, fungus, scanning Kelvin probe
1. Introduction Microbially influenced corrosion (MIC) refers to the metal deterioration process influenced by the presence and/or the metabolic activities of microorganisms (Ref 1). It has been widely found in different natural and industrial environments and can contribute to a significant portion of corrosion-induced economic loss. According to an estimate in 1993, the direct cost of MIC in the United States fell in the range of $30-$50 billion/ year (Ref 2). Both prokaryotic (i.e., bacteria, archaea) and eukaryotic (i.e., fungi) microorganisms can cause MIC (Ref 3). To date, considerable interests have been given to corrosion influenced by prokaryotic bacteria cells. A well-known example is sulfatereducing bacteria (SRB), which are anerobic and live predominantly in environments such as soils (Ref 4-6) and seawater (Ref 7, 8). To our best knowledge, the accelerating or inhibiting effect of fungus to corrosion has not been adequately investigated. Most fungi are aerobic and reproduce most favorably in humid aerobic environments (Ref 9-11). They have been long identified as serious threats which cause corrosion of materials in
Electronic supplementary materialThe online version of this article (doi:10.1007/s11665-015-1558-2) contains supplementary material, which is available to authorized users. Dawei Zhang, Feichi Zhou, Kui Xiao, Tianyu Cui, Hongchong Qian, and Xiaogang Li, Corrosion & Protection Center, University of Science & Technology Beijing, Fushi Bldg., 30 Xueyuan Rd., Haidian District, Beijing 100083, China. Contact e-mail:
[email protected].
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various industries. For example, the widespread corrosion problems of aluminum aircraft fuel tanks have been attributed to acid-producing fungi, most notably Hormoconis resinae (Ref 12). It has also been shown that fungi such as Aspergillus niger, Candida mycoderma, and Saccharomyces cerevisiae can increase the risks of corrosion of food contacting equipment (Ref 13). In a 2-year laboratory test under humid atmosphere, A. niger was found to accelerate zinc corrosion due to the diminished thickness of the protective inner layer of corrosion products, whereas it inhibited aluminum corrosion by promoting passivity at the sites of localized corrosion (Ref 14). The mechanisms by which microorganisms influence corrosion are complicated. On one hand, they produce metabolites including acids, sulfide, ammonia, and other corrosive species which can accelerate the electrochemical reactions of corrosion (Ref 3). The heterogeneous surface colonization of microorganisms can also increase the risks of localized corrosion such as pitting (Ref 15), stress corrosion cracking (Ref 16), and hydrogen embrittlement (Ref 17). On the other hand, microorganisms can inhibit corrosion by neutralizing corrosive species during their metabolisms and forming protective biofilms over metal substrates (Ref 18). The MIC mechanisms can be even more complex when multiple species of microorganisms exist and interact with each other. For example, precipitates by ironoxidizing bacteria (IOB) on 316L stainless steel can form an oxygen-limiting environment which promoted the growth of SRB, thereby resulting in a higher corrosion rate than in the presence of only SRB or IOB (Ref 19). Most of the current studies focus on MIC in immersed condition, where the formation of biofilms is relatively fast. Few have investigated the influence of microorganisms on metal corrosion under atmospheric environment (Ref 14). In this study, 304 stainless steel and titanium were exposed to two very common fungi, Paecilomyces variotii and A. niger, as well as their mixture in highly humid atmosphere. The fungus proliferation on each type of material was observed via
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scanning electron microscopy (SEM). By scanning Kelvin probe (SKP), the individual effect of each fungus and the synergistic effect of their mixture on the initial corrosion behaviors (up to 28 days) of 304 stainless steel and titanium were compared. Long-term exposures for 60 days were also carried out and the corrosion behaviors were evaluated based on stereomicroscopic observation.
2. Experimental 2.1 Materials 304 stainless steel and titanium specimens (10 mm 9 10 mm 9 4 mm) were sequentially polished with 240, 400, 800, 1200, and 2000 grit sandpapers, followed by cleaning with acetone and sterilized with ethanol. All chemicals were purchased from SinoPharm and used without further purification. The distilled water and all glassware were autoclaved at 120 °C for 20 min, followed by UV sterilization for 30 min in a cell culture hood (HDL Class II). P. variotii and A. niger suspension were obtained from Institute of Microbiology, Chinese Academy of Sciences. The concentration of the fungus suspensions are 1 9 107 spores/L.
2.2 Fungus Corrosion Test The nutrient solution was prepared by dissolving the following salts in 1 L distilled water: KH2PO4 (0.7 g), K2HPO4 (0.7 g), MgSO4Æ7H2O (0.7 g), NH4NO3 (1.0 g), NaCl (0.005 g), FeSO4Æ7H2O (0.002 g), ZnSO4Æ7H2O (0.002 g), and MnSO4ÆH2O (0.001 g). The pH of the obtained solution was 6.5. Prior to use, the solution was sterilized with UV lights for 30 min.
For the fungus corrosion test, the spore suspension was diluted in the nutrient solution (1:9 v/v) and sprayed on the specimen surface. Subsequently, the specimens were transferred to an incubator and stored at 30 °C and 95% relative humidity (RH). After 7, 14, 21, and 28 days, the specimens were retrieved and the surface morphology was observed under scanning electron microscope (SEM, FEI Quanta 250). To study the corrosion behaviors, distribution of Kelvin potential on the specimen surface was mapped by SKP measurements using a PAR M370 electrochemistry workstation. Calibrated at a distance of 100 lm from the specimen surface, the probe was vibrating at an amplitude of 30 lm and a frequency of 80 Hz. Step scan was then performed over an area of 2000 lm 9 2000 lm. All measurements were carried out at ambient temperature and 60% RH. To study the long-term corrosion behaviors in the presence of fungus, the specimens were sprayed with fungus suspension and incubated at 30 °C and 95% RH for 60 days. The surface morphology of the specimen was then observed by SEM and stereomicroscopy (KENYENCE VHX-2000).
3. Results and Discussion 3.1 Surface Morphology Figure 1 presents the surface morphologies of 304 stainless steel exposed to P. variotii suspension. After 7 days of exposure at 30 °C and 95% RH (Fig. 1a), a small number of spores are found at specimen surface, with the size of 2-10 lm. From 14 to 28 days (Fig. 1b-d), spores start to form mycelia, which continue to grow and cover an increasing area. Corrosion products are clearly visible surrounding the spores after
Fig. 1 Growth of P. variotii on 304 stainless steel after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days. Scale bar is 50 lm. Corrosion products are marked with arrows
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Fig. 2
Growth of P. variotii on titanium after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days. Scale bar is 50 lm
Fig. 3
Growth of A. niger on 304 stainless steel after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days. Scale bar is 50 lm
21 days of exposure. At 28 days, conidiophorous filaments are observed, indicating that the fungus proliferation was stable and the new generations were produced. In the control group where only nutrient solution is present (Fig. S1), no corrosion product was observed after 28 days. Energy-dispersive x-ray spectroscopy (EDS) analyses were performed to reveal the compo-
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sitional differences on the specimen surface (Fig. S2). The bare surface shows an atomic ratio of Fe-O close to 28:3. Covered by corrosion products, the region around the spores exhibits marked increases in oxygen (Fe:O 2:1) and carbon (Fe:C 3:1) contents. This carbon enrichment effect has been previously observed in the corrosion product of zinc after fungus
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Fig. 4
Growth of A. niger on titanium after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days. Scale bar is 50 lm
Fig. 5
Growth of mixed strains on 304 stainless steel after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days. Scale bar is 50 lm
exposure (Ref 14). It was attributed to the well-known acidforming ability of fungus which can secrete a variety of organic metabolites such as oxalic, acetic, lactic, and citric acids (Ref 20). It is expected that these acids can accelerate the cathodic reaction and thereby the overall corrosion process (Eq 1 and 2) (Ref 21, 22):
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Anodic reaction: 1=2Fe ! 1=2Fe2þ þ e ;
Fe2þ ! Fe3þ þ e ;
ðEq 1Þ
Cathodic reaction 1=4O2 þ 1=2H2 O þ e ! OH :
ðEq 2Þ
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Fig. 6
Growth of mixed strains on titanium after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days. Scale bar is 50 lm
Fig. 7 Potential distribution on 304 stainless steel exposed to P. variotii after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days and (e) the Gaussian fitting curves (Color figure online)
Compared to 304 stainless steel, the growth of P. variotii is slower on titanium surface, as indicated by the shorter mycelia (Fig. 2). During the entire exposure period, no corrosion product was found on the specimen surface. Titanium has been long noted for its superior corrosion resistance thanks to a compact titanium oxide layer on its surface. Thus, it is less likely to produce metallic ions as a result of corrosion which serve as nutrients sustaining the metabolic activities of microorganisms (Ref 23-25). On 304 stainless steel, a slightly slower growth rate is observed for A. niger than P. variotii, as seen from the shorter
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mycelia (Fig. 3). Unlike P. variotii which existed as isolated spherical spores, A. niger formed clusters and grew in a radial fashion. From EDS analysis, the enrichment of oxygen and carbon in the corrosion products near the spores also suggests that the corrosion process was accelerated due to the acids from fungus metabolisms (Fig. S3). Figure 4 shows the proliferation of A. niger on titanium, which appears to be similar to the case of 304 stainless steel. No corrosion product can be found after 28 days of exposure. When mixed strains of P. variotii and A. niger were cocultured on the specimen surfaces, a more rapid fungus
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Fig. 8 Potential distribution on titanium exposed to P. variotii after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days and (e) the Gaussian fitting curves (Color figure online)
Fig. 9 Potential distribution on 304 stainless steel exposed to A. niger after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days and (e) the Gaussian fitting curves (Color figure online)
proliferation can be observed on both 304 stainless steel (Fig. 5) and titanium (Fig. 6). This may be explained by that the organic acids produced by P. variotii can be utilized as carbon sources for the growth of A. niger (Ref 26, 27). Despite the increase in fungus proliferation, not much corrosion product was observed on both 304 stainless steel and titanium, possibly due to the consumption of oxygen in the vicinity of fungus growth (Ref 18).
3.2 SKP analysis In most MIC studies, corrosion behaviors were evaluated by macroscopic methods such as polarization scans (Ref 13, 19) or
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electrochemical impedance spectroscopy (EIS) (Ref 14, 16) which reflect only average corrosion information at the entire electrode/electrolyte interface. Instead, microscopic electrochemical technique such as scanning vibrating electrode technique (SVET) (Ref 28) and wire beam electrode (WBE) (Ref 5, 29, 30) have been more recently proposed as better tools for MIC studies, considering that MIC often occurs more locally as a result of heterogeneous microbial attachment and colonization. Nevertheless, all of these methods are limited to measurements under aqueous environments and are not suitable for evaluation of fungus corrosion in atmosphere. Operated under thin electrolyte in air, SKP has a unique capability of providing highly sensitive and non-destructive measurements
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Fig. 10 Potential distribution on titanium exposed to A. niger after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days and (e) the Gaussian fitting curves (Color figure online)
Fig. 11 Potential distribution on 304 stainless steel exposed to mixed strains after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days and (e) the Gaussian fitting curves (Color figure online)
for atmospheric corrosion in a finite area (Ref 31). It measures the difference between work functions of working electrode and reference electrode (Kelvin probe), thereby revealing the condition and electrochemical activity of the target surface (Ref 32, 33). Thus, it is an ideal tool for capturing the localized potential differences during the initial stages of fungus corrosion. However, to our best knowledge, the use of SKP in MIC studies has been seldom reported (Ref 34). In this study, SKP analyses were performed over an area of 2000 lm 9 2000 lm on specimens after different exposing periods. Figure 7 shows the potential distribution of 304 stainless steel surface exposed to P. variotii. The surface potential data were also fitted with Gauss formula (Eq 3, Ref 35) and the results are plotted in Fig. S4 and 7(e):
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" # A 2ðx lÞ2 ; y ¼ y0 þ pffiffiffiffiffiffiffiffi exp r2 r p=2
ðEq 3Þ
where y0 is the ordinate offset; A is a constant; l is the expectation value of Gaussian distribution and the location of the peak value of Kelvin potential; r2 is the variance of Gaussian distribution which measures the width of Kelvin potential distribution. Initially, the surface of 304 stainless steel presents a relatively uniform potential distribution with peak value located at 0.9 V (Fig. S4). After 7 days, peak value of Kelvin potential was increased to 0.67 V. Considering the minimal fungus growth at this stage (Fig. 7a), this increase was predominantly abiotic and can be attributed to the
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Fig. 12 Potential distribution on titanium exposed to mixed strains after (a) 7 days; (b) 14 days; (c) 21 days; and (d) 28 days and (e) the Gaussian fitting curves (Color figure online)
Fig. 13 Growth of (a) P. variotii, (b) A. niger, and (c) mixed strains on 304 stainless steel after 60 days and (d-f) the corresponding surface morphologies by stereomicroscopy. Scale bar is 50 lm (Color figure online)
rapid surface oxidation of stainless steel in the humid atmospheric environment (Ref 36). As the growth of P. variotii continued, the surface potential of stainless steel experienced a further increase and eventually reached to 0.35 V after 28 days. Such ennoblement effect has been extensively reported in many previous MIC studies of passive alloys, especially stainless steels (Ref 37). Although the clear causes remain controversial, this phenomena is generally attributed to the increase in the cathodic reaction rate by microbial metabolic activities (Ref 28, 37-39). In addition, the non-uni-
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form colonization of microorganisms and the formation of corrosion products have also led to a more heterogeneous potential distribution which may increase the driving force for localized corrosion (Ref 37, 40, 41). The heterogeneous potential distribution can also be confirmed by the widened Gaussian fitting curve in Fig. 7(e). On titanium surface, the initial 7 days exposure results in a large increase in the peak value of Kelvin potential from 0.9 to 0.53 V (Fig. 8e). Combined with the surface morphology shown in Fig. 2, such increase can be attributed to the
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formation of compact titanium oxide layer in the humid atmospheric environment. The smaller increase in potential upon further exposure and the narrower potential distribution after 28 days both indicate a better resistance of titanium to fungus corrosion than 304 stainless steel. The SKP analyses of 304 stainless steel and titanium exposed to A. niger are shown in Fig. 9 and 10, respectively. For 304 stainless steel, the peak value of Kelvin potential was raised to 0.53 V after 28 days, which was much lower than the specimen exposed to P. variotii (0.35 V) (Fig. 7). In addition, a relatively uniform potential distribution is obtained (Fig. 9e). As shown in Fig. 10, similarly, exposure to A. niger results in a smaller increase in surface potential of titanium from 7 d to 28 days as well as a more uniform potential distribution at 28 days. Thus, it can be concluded that A. niger was less corrosive to both 304 stainless steel and titanium compared with P. variotii. Figure 11 shows the potential distribution on 304 stainless steel surface exposed to a mixture of P. variotii and A. niger. The results correlated well with SEM observation as shown in Fig. 5. From 7 to 28 days, the peak values of Kelvin potential are located in a narrow range from 0.6 to 0.4 V and only a small increase is observed. Past research on corrosion in presence of mixed populations of microorganisms have mostly demonstrated a synergistic effect by which corrosion was accelerated (Ref 42). In a number of studies, mixed microorganisms were also found to exert an inhibitive effect toward corrosion by forming more protective corrosion products (Ref 7, 43) or compact biofilms (Ref 44). In our case, the faster fungus proliferation did not concurrently lead to an aggravated corrosion. This may be explained by the increased oxygen consumption from faster fungus proliferation. For example, Jayaraman et al. have demonstrated metabolic activities of aerobic microorganisms such as Pseudomonas fragi can cause
oxygen depletion at metal/biofilm interface which significantly inhibited the corrosion rate of mild steel (Ref 45, 46). Exposing titanium surface to the mixture of P. variotii and A. niger only caused a slight corrosion (Fig. 12). After 28 days of exposure to the mixed strains, the peak potential values of both 304 stainless steel (Fig. 11) and titanium (Fig. 12) are similar to the cases where only A. niger was present (Fig. 9 and 10). This suggests that A. niger might be the dominating strain in MIC on these metals.
3.3 Long-Term Exposure Fungus exposures were also carried out in atmospheric environment at 30 °C and 95% RH for 60 days in order to study the long-term effects of P. variotii and A. niger on corrosion behaviors of 304 stainless steel and titanium. On the surface of stainless steel, P. variotii have grown into highly clusterous colonies after 60 days (Fig. 13a), whereas A. niger are sporadically distributed in the form of long mycelia (Fig. 13b). For both fungi, corrosion products can be found in the surrounding area. As shown in Fig. 13(c), the mixture of two fungi resulted in the most severe fungus proliferation. To compare the corrosion morphologies for each specimen, stereomicroscopy was used for direct observation of the three-dimensional profiles of specimen where fungi had been gently removed. Deeper and larger corrosion grooves were seen on the specimen exposed to P. variotii while the grooves are shallower and more scattered in the case of A. niger. The specimen exposed to the mixed strains was least corroded, which corresponded well with results after short-term exposures. Compared to 304 stainless steel, titanium surface exhibits a better resistance to fungus growth (Fig. 14). From stereomicroscopic observation, only a small amount of corrosion product can be found in the regions closed to fungus growth after 60 days of exposure.
Fig. 14 Growth of (a) P. variotii, (b) A. niger, and (c) mixed strains on titanium after 60 days and (d-f) the corresponding surface morphologies by stereomicroscopy. Scale bar is 50 lm (Color figure online)
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4. Conclusions In summary, we have studied the initial corrosion behaviors of 304 stainless steel and titanium in 28 day exposures to P. variotii and A. niger in highly humid atmospheric environment. After a long-term exposure for 60 days, fungus proliferation and corrosion morphologies were also evaluated. Based on the results, the following conclusions can be drawn: 1. The SEM studies showed a higher proliferation rate of P. variotii than A. niger and the fastest proliferation was observed in the case of the mixed strains. Compared to 304 stainless steel, titanium surface exhibited a better resistance to fungus growth. 2. Compared to A. niger, P. variotii has resulted in a more serious corrosion as indicated by the larger increase in surface potentials as well as more heterogeneous potential distributions in SKP analyses. 304 stainless steel was more susceptible to attacks from fungus corrosion than titanium. 3. Despite the fastest fungus growth, only a mild corrosion was observed in the presence of the mixed strains, possibly due to the higher oxygen consumption by increased fungus proliferation.
Acknowledgments This work is supported by National Natural Science Foundation of China (No. 51401018), the National Basic Research Program of China (973 Program Project, No. 2014CB643300).
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41. F.N. Afshar, J. de Wit, H. Terryn, and J. Mol, Scanning Kelvin Probe Force Microscopy as a Means of Predicting the Electrochemical Characteristics of the Surface of a Modified AA4xxx/AA3xxx (Al Alloys) Brazing Sheet, Electrochim. Acta, 2013, 88, p 330–339 42. I.B. Beech and J. Sunner, Biocorrosion: Towards Understanding Interactions Between Biofilms and Metals, Curr. Opin. Biotechnol., 2004, 15, p 181–186 43. Y. Duan, S.-M. Li, J. Du, and J.-H. Liu, Corrosion Behavior of Q235 Steel in the Presence of Pseudomonas and Iron Bacteria, Acta Phys. Chim. Sin., 2010, 26, p 3203–3211 44. N.O. San, H. Nazır, and G. Do¨nmez, Evaluation of Microbiologically Influenced Corrosion Inhibition on Ni-Co Alloy Coatings by Aeromonas salmonicida and Clavibacter michiganensis, Corros. Sci., 2012, 65, p 113–118 45. A. Jayaraman, E.T. Cheng, J.C. Earthman, and T.K. Wood, Importance of Biofilm Formation for Corrosion Inhibition of SAE 1018 Steel by Axenic Aerobic Biofilms, J. Ind. Microbiol. Biotechnol., 1997, 18, p 396–401 46. A. Jayaraman, J.C. Earthman, and T.K. Wood, Corrosion Inhibition by Aerobic Biofilms on SAE 1018 Steel, Appl. Microbiol. Biotechnol., 1997, 47, p 62–68
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