Enzyme-driven mechanisms in biocorrosion

Objectives (abstract of presentation): Recent works carried out in our team concerning enzymes and biocorrosion are presented at the meeting. For aerobic conditions, the direct catalysis of the reduction of oxygen on steel by enzymes or porphyrin was proved and a local electrochemical analysis technique (SVET) was developed to visualize the localization of the catalysis. On anaerobic conditions, the influence of phosphate species and other weak acids on the water reduction on steel was shown. The role of hydrogenase in biocorrosion process was also studied. Any conclusion: In aerobic conditions, many complementary contributions including enzymatic catalysis exist and can reproduce biocorrosion in the lab (MOB, GOD, hopefully porphyrin…) and even more inter-penetration of the thematic areas is still required. In anaerobic conditions, revisiting the previous lab studies taking into account possible involvement of phosphate species (common buffer) is certainly necessary and other mechanism than the FeS formation can also explain biocorrosion as the direct electron transfer between hydrogenase and steel material

Membrane electrochemical reactors (MER) for NADH regeneration in HLADH-catalysed synthesis: comparison of effectiveness

Two membrane electrochemical reactors(MER) were designed and applied to HLADH-catalysed reduction of cyclohexanone to cyclohexanol. The regeneration of the cofactor NADH was ensured electrochemically, using either methyl viologen or a rhodium complex as electrochemical mediator. A semipermeable membrane (dialysis or ultra-filtration) was integrated in the filter-press electrochemical reactor to confine the enzyme(s) as close as possible to the electrode surface. When methyl viologen was used, the transformation ratio of cyclohexanone varied from 0 to 65% depending on the internal arrangement of the reactor. Matching the reactor configuration to the reaction system was essential in this case. With the rhodium complex, the ultra-filtration MER was tested in continuous and recycling configurations. The best conditions led to 100% transformation of 0.1 L volume of 0.1 M cyclohexanone after 70 h with the recycling mode. Finally, the performances of the reactors are discussed with respect to different evaluations of the production yields

Study of the influence of phosphoric acid in steel anaerobic corrosion via the hydrogen evolution reaction.

Although it is well known that the phosphate species are quiet electrochemically inert when considering their calculated redox potential, few studies since the end of 80’s reported electrochemical phenomenon observed in the reduction domain which were linked to the presence of phosphate species. Some authors proposed that phosphate ions and weak acids in general catalysed the electrochemical reduction of water [1-3] following the kind of reactions written above: Electrochemical reactions H2PO4- + e- Had + HPO42- (1) Had + H2PO4- + e- H2 + HPO42- (2) Acido-basic equilibrium H2PO4- + OH-- H2O + HPO42- (3) Combining the reactions, the reduction of water was obtained 2 H2O + 2e- H2 + 2OH- (4) Marinovic and Despic [1] assumed that the hydrogen evolution was more likely to occur from species other than water, since the bond between the hydrogen atom and the rest of the molecule is weaker in weak acid compared with the hydrogen-oxygen bond in a water molecule. This was shown on silver electrodes [2] and also on platinum ones [1]. In a previous work [3], we showed that phosphate ions exhibited the same electrochemical behaviour on stainless steel. A theoretical model was proposed according to the experimental data the model showed that a significant quantity of molecular hydrogen was produced by the mechanism presented above. In the present paper, the influence of phosphoric acid on hydrogen evolution reaction has been studied by voltammetry on platinum and stainless steel rotating disc electrodes. A linear correlation between acid concentration and the absolute value of the current at certain reduction potential has been verified. The effect of the pH value of the solution in the obtained current has demonstrated to be strongly related to the dissociation state of the acid. In a second part, as the hydrogen evolution reaction is the principal cathodic reaction of metal corrosion in anaerobic aqueous environments, the role that weak acids can play in this process was studied. Corrosion experiments on carbon steel were carried out in phosphoric acid solution under anaerobic conditions. The influence of hydrogen in the solution was also studied. In the absence of oxygen, a substantial increase of corrosion rate was found when carbon steel samples were submerged in low phosphoric acid concentration solution compared to that obtained in distilled water

Geobacter sulfurreducens can protect 304L stainless steel against pitting in conditions of low electron acceptor concentrations

The effect of Geobacter sulfurreducens cells was studied on the electrochemical behaviour of 304L stainless steel, emphasizing the role of the soluble electron acceptor (fumarate). In fumarate-lacking media, the presence of G. sulfurreducens induced free potential ennoblement in a few hours. This ennoblement has already been observed in standard media that contained fumarate. Our previous studies have shown that G. sulfurreducens shifted the pitting potential toward the positive values. The pits induced by the presence of the bacteria were wider and deeper than in the absence of bacteria. Here, in fumarate-lacking media, similar shift in pitting potential was observed, but the repassivation phase was strongly improved. AFM analysis showed that pits were identical with those observed in the absence of bacteria at lower potential. In contrast with all the previous work where G. sulfurreducens enhanced corrosion, here at a low concentration of electron acceptor, the presence of the bacteria protected the steel against pitting

Effect of the semi-conductive properties of the passive layer on the current provided by stainless steel microbial cathodes

Geobacter sulfur reducens biofilms were formed under constant polarisation at −0.6 V vs. Ag/AgCl on stainless steel cathodes to catalyse the reduction of fumarate. The time-evolution of the current strongly depended on the quality of the inoculum. Inoculating with young cells significantly shortened the initial lag-phase and using the same inoculum improved the reproducibility of the current–time curves. The whole set of experiments showed that 254SMO stainless steel provided higher current densities (on average 14.1 A/m2) than biofilms formed on 316L stainless steel (on average 4.5 A/m2). Biofilm coverage assessed by epifluorescent microscopy showed that coverage ratios were generally higher for 316L than for 254SMO. It must be concluded that 254SMO is more efficient in transferring electrons to bacterial cells than 316L. Mott–Schottky diagrams recorded on both materials under conditions of electrolysis in the absence of microorganisms showed that the surface oxide layers had similar n-type semi-conductive behaviour for potential values higher than the flat band potential. In contrast, 316L exhibited slight p-type behaviour at potential lower than the flat band potential, while 254SMO did not. The higher electrochemical performances of biocathodes formed on 254SMO are explained by semi-conductive properties of its passive layer, which prevented the p-type behaviour occurring in cathodic electrolysis conditions

Possible role of Geobacter sulfurreducens in anaerobic corrosion of steels

Geobacteraceae are the most widespread microorganisms in soils and sediments in which microbial reduction of Fe(III) is an important process, either in the natural degradation of organic compounds or in their bioremediation. Geobacter species have been shown to be predominant microorganisms on electrodes harvesting electricity from the sediments. They have the capability to oxidize organic electron donor to carbon dioxide transferring the electron directly to electrodes [1]. On the other side, the ability of Geobacter sulfurreducens to reduce nitrate to nitrite or fumarate to succinate with a graphite electrode serving as electron donor has also been demonstrated [2]. Direct electron transfer to solid electrodes is achieved through periplasmic and outer membrane c-type cytochromes [3]. Outer membranes proteins and even some kind of conductive pili that serve as biological nanowires are also involved in the electron transfer chains, mainly to Fe(III) and Mn(IV) oxides [4]. The aim of this study was to assess the possible influence of G. sulfurreducens on the occurrence of corrosion of steels. Experiments were performed with pure cultures of G. sulfurreducens on mild steel (XC45) and three different kinds of stainless steels (ferritic steel, 304L, 316L). In each case the free potential increased by 200 to 300 mV after the injection of the bacteria. On the contrary, control experiments performed with the injection of the sterile medium or the bacteria suspension after filtration on a 0.2 µm filter did not induce any variation in the free potential. The presence of the cell was consequently directly responsible for the potential increase of the coupons. The occurrence or not of corrosion was discussed with respect to this potential increase and the nature of the medium. Besides, preliminary results allow assessing the possibility to use G. sulfurreducens to design protective biofilms

Local analysis of oxygen reduction catalysis by scanning vibrating electrode technique : a new approach to the study of biocorrosion

The scanning vibrating electrode technique (SVET)was employed to investigate oxygen reduction catalysis by the presence of enzyme in an aerobic medium. Heme protoporphyrin (hemin) was chosen as a model of the enzymes that are able to catalyze oxygen reduction. A strict experimental protocol was defined for preparing the graphite surface by deposition of hemin with a simple configuration mimicking the presence of enzyme on the samples. The same configuration was adapted to a stainless steel electrode. Different geometric arrangementswere investigated by SVET to approach the local conditions. The results demonstrated that hemin deposited on the electrode surface led to an increase in the cathodic current, which indicated a catalytic effect. Based on the SVET analysis, itwas demonstrated that hemin caused the appearance of galvanic cells on the material surface. The SVET proved able to locate active catalytic centres and therefore to foresee the contribution of the enzyme to the creation of galvanic cells, thus leading to localized corrosion. The application of SVET to the study of the interaction between biological molecules and material provides a newapproach for visualizing and understanding microbially influenced corrosion (MIC) in an aerobic medium

Effect of hydrogenase on the corrosion of mild steel

Losses due to corrosion are evaluated at 4% of the GDP of industrialised countries and biocorrosion may be responsible for 10% of these costs [1]. Whereas the general mechanism of anaerobic corrosion, involving iron sulphur deposits, seems now well agreed, the detailed mechanism is still unclear and the implication of hydrogenase is very controversial. The influence of a [Fe] hydrogenase from C. acetobutylicum on mild steel corrosion was studied using a galvanic cell and measuring the current and the free potential. This hydrogenase seems to induce pitting corrosion. The mechanism is discussed using activated and deactivated enzyme and the possible influence of phosphate is highlighted. In the galvanic cell, the presence of hydrogenase on the surface of only one electrode induced a galvanic current up to 10µA and the potential decreased by 500mV in the presence of phosphate. MEB and EDX analysis confirmed the presence of a vivianite deposit which forms a protective barrier to corrosion [2]. It seems that either type of hydrogenase ([Fe] or [Ni-Fe]) has a significant role in initiating corrosion [3, 4]

Effect of Geobacter sulfurreducens on the microbial corrosion of mild steel, ferritic and austenitic stainless steels

The influence of Geobacter sulfurreducens was tested on the anaerobic corrosion of four different steels: mild steel 1145, ferritic steel 403 and austenitic steels 304L and 316L. Within a few hours, the presence of cells induced a free potential (Eoc) ennoblement around +0.3 V on 1145 mild steel, 403 ferritic steel and 304L austenitic steels and slightly less on 316L. The kinetics of Eoc ennoblement depended on the amount of bacteria in the inoculum, but the final potential value depended essentially on the nature of the material. This effect was due to the capacity of G. sulfurreducens to create a direct cathodic reaction on steel surfaces, extracting the electrons directly from material. The presence of bacterial cells modified the corrosion features of mild steel and ferritic steel, so that corrosion attacks were gathered in determined zones of the surface. Local corrosion was significantly enhanced on ferritic steel. Potential ennoblement was not sufficient to induce corrosion on austenitic steels. In contrast G. sulfurreducens delayed the occurrence of pitting on 304L steel because of its capability to oxidize acetate at high potential values. The electrochemical behaviour of 304L steel was not affected by the concentration of soluble electron donor (acetate, 1–10 mM) or the amount of planktonic cells; it was directly linked to the biofilm coverage. After polarization pitting curves had been recorded, microscopic observations showed that pits propagated only in the surface zones where cell settlement was the densest. The study evidenced that Geobacter sulfurreducens can control the electrochemical behaviour of steels in complex ways that can lead to severe corrosion. As Geobacteraceae are ubiquitous species in sediments and soils they should now be considered as possible crucial actors in the microbial corrosion of buried equipment