effect of mixed species biofilm on corrosion of cast iron
TRANSCRIPT
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• By • Farhad Batmanghelich
• Youngwoo Seo
• Department of Chemical and Environmental Engineering
• University of Toledo
September 2015
Effect of Mixed Denitrifying and Sulfate Reducing Bacterial
Biofilms on Corrosion Behavior of Cast Iron
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Outline
2
Introduction and Theoretical background
Scope of Study Materials and
Methods Results and Discussions
Conclusions
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Outline
3
Introduction and Theoretical background
Scope of Study Materials and
Methods Results and Discussions
Conclusions
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Corrosion
4
Corrosion is the deterioration of a material due to the reactions with its surrounding environment
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5
Corrosion Economic Impact
Utilities
47.9 billion $
Production and
manufacturing
17.6 billion $
Government
20.1 billion $
Infrastructure
22.6 billion $
Transportation
29.7 billion $
b 43
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Why knowing about corrosion mechanisms is
important?
6
Utilities
47.9 billion $
Production and
manufacturing
17.6 billion $
Government
20.1 billion $
Infrastructure
22.6 billion $
Transportation
29.7 billion $
b 43
Corrosion mitigation strategies:
1- Mechanism identification
2- Occlude those mechanisms
3- Monitor
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Corrosion in Water Systems
7
Decrease in water capacity………(Energy Loss)
Holes and leakages………………(Structural Integrity)
Drinking water quality…………...(Health Problems)
Red water………………………...(Aesthetic problems)
Odor……………………………...(Aesthetic Problem)
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The Cycle of Corrosion
8
Ore
Thermodynamically
stable state Energy
Reduction
Metals &
Alloys Products
Corrosion
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Electrochemistry
Corrosion is an interfacial electrochemical process
Electrochemical rxns are redox rxns in which:
Electron transfer over extremely large distances
In a redox rxn oxidation state of the reactants change
9
Interface (Double
layer)
Bulk
Surf
ace
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Electrochemistry
10
Full reaction Half reactions
(electrode reactions)
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Electrochemistry
11
Anodic half
reaction
Cathodic half
reaction
Equilibrium Thermodynamics (Nernst equation)
Non-equilibtium Kinetics (Butler-Volmer; Conttrell, etc.)
The rate of an electrochemical reaction is expressed
in terms of electric current.
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Electrochemical Techniques
12
Basis of all electrochemical techniques:
A deliberate perturbation in one of the parameters of the system and
monitoring the response of the system.
(a) Potential Step : { chronoamperometry, chronopotentiometry, chronocoulometry, etc.}
(b) Potential sweep : { linear scan voltammetry, cyclic voltametry, etc.}
Potentiodynamic polarization
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Electrochemical Impedance Spectroscopy
(EIS)
Electrochemical Impedance Spectroscopy (EIS)
13
Nyquist Bode
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Electrochemical Impedance Spectroscopy
(EIS)
14
The sine AC energy can be either stored or dissipated
at the interface by different layers.
The interface of a metal and its surroundings can
be modeled based on physical geometry and properties
of the surface.
The model can then be fitted to the impedance data
to acquire physical values such as interfacial resistance
and capacitances.
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Biocorrosion (the role of biofilm)
Any material or phenomena that
changes the interface of a metal and
its environment is able to affect
electrochemical reactions, thence
alters corrosion behavior.
15
Biofilms form on metal surfaces
Biofilms change corrosion behavior
Biofilms can either
exacerbate or alleviate
corrosion hazard.
In studying corrosion;
effect of bulk
metabolites and biofilms
should be considered
separately.
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Sulfate Reducing Bacteria (SRB)
SRB species such as Desulfovibrio Vulgaris reduce
SO42-
to S2- and eventually produce H2S.
H2S is corrosive….
SRB are anaerobes.
16
Venzlaff, corr.sci, 66, 2013
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Pseudomonas aeruginosa
Pseudomonas aeruginosa (PAO1) is an opportunistic pathogen with a high level of
adaptability to various environments.
It is a facultative bacteria and can survive in both aerobic and anaerobic environments
Under anaerobic conditions it can respire through denitrification
Denitrification:
Reduction of nitrate (NO3-) to nitrite (NO2
-), ammonia and N2.
17
PAO1 wild type
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Mixed Species Corrosion
Bacteria usually live in consortia.
Denitrifying and Sulfate Reducing Bacteria (SRB) are abundant in aqueous environments.
Under corrosion tubercles there is no oxygen, therefore PAO1 and SRB can exist together
in that area.
Mixed species consortia can exacerbate or alleviate corrosion.
18
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19
Introduction and Theoretical background
Scope of Study Materials and
Methods Results and Discussions
Conclusions
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Scope of Study
Main objective of this study is to evaluate effect of mixed bacterial biofilms and cultures
on corrosion behavior of cast iron.
(i) What impact mixed model bacteria solutions containing corresponding metabolites have
on corrosion of cast iron?
(ii) How biofilm composition and morphology contributes to corrosion of cast iron in the
absence of bacterial metabolites?
20
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Outline
21
Introduction and Theoretical background
Scope of Study Materials and
Methods Results and Discussions
Conclusions
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Materials and Methods, Bacteria Culture
Preparation
D. vulgaris and P.aeruginosa were
grown anaerobically in an anaerobic
chamber. 95% N2- 5% H2
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Materials and Methods, Bacteria Culture
Preparation
D. vulgaris and P.aeruginosa were grown
anaerobically in an anaerobic chamber. 95%
N2- 5% H2
All the seed preparations were made in a sterile
clean hood under UV light with ethanol
sterilization. Coupons were also sterilized by
70% ethanol and subsequent UV irradiation.
23
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Coupon Preparation
24
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PAO1 and SRB seeds
SRB produces Hydrogen Sulfide (H2S). Iron reacts with H2S to
form black FeS precipitates. A procedure for checking SRB
growth is to add iron salts to grown SRB cultures to see if black
precipitates form.
25
PAO1 SRB
Anaerobic Hungate
tubes
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Overview of experiments
26
In-situ electrochemical
experiments
• EIS and potentiosynamic polarization experiments on coupons that have been immersed for 1, 3 and 7 days in bacterial solutions of PAO1, SRB and mixed species.
• Corrosion weight loss : 1, 3 and 7 days immersion bacteria cultures; ASTM D-2688.
Ex-situ electrochemical
experiments
• EIS and potentiodynamic polarization tests in 3.5% NaCl solution (pH = 4), on coupons that have been immersed in bacterial solutions of PAO1, SRB and mixed for 1, 3 and 7 days.
• Corrosion weight loss : 1, 3 and 7 days immersion in bacteria + immersion in 3.5% NaCl solution (pH = 4) for 4 days; ASTM D-2688.
Biofilm quantification
and observation
• EPS extraction [Cationic Exchange Resin (CER) method] and quantification [phenol sulfuric acid (PSA) for polysaccharides; Coomassie blue method for proteins].
• Fluorescence microscopy
• Atomic Force Microscopy (AFM)
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Corrosion Intrumentation (ex-situ)
27
For in-situ electrochemical experiments: A multiportTM
Corrosion Cell kit (Gamry) connected to a Reference 600
potentiostat (Gamry) was used.
For ex-situ electrochemical experiments: A cell was
designed to maintain anaerobiosis during the course of ex-
situ eperiments.
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Corrosion Intrumentation (in-situ)
28
For in-situ electrochemical experiments: A multiportTM
Corrosion Cell kit (Gamry) connected to a Reference 600
potentiostat (Gamry) was used.
For ex-situ electrochemical experiments: A cell was
designed to maintain anaerobiosis during the course of ex-
situ eperiments.
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Outline
29
Introduction and Theoretical background
Scope of Study Materials and
Methods Results and Discussions
Conclusions
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Results and Discussion
30
In-situ electrochemical
experiments
• EIS and potentiosynamic polarization experiments on coupons that have been immersed for 1, 3 and 7 days in bacterial solutions of PAO1, SRB and mixed species.
• Corrosion weight loss : 1, 3 and 7 days immersion bacteria cultures; ASTM D-2688.
Ex-situ electrochemical
experiments
• EIS and potentiodynamic polarization tests in 3.5% NaCl solution (pH = 4), on coupons that have been immersed in bacterial solutions of PAO1, SRB and mixed for 1, 3 and 7 days.
• Corrosion weight loss : 1, 3 and 7 days immersion in bacteria + immersion in 3.5% NaCl solution (pH = 4) for 4 days; ASTM D-2688.
Biofilm quantification
and observation
• EPS extraction [Cationic Exchange Resin (CER) method] and quantification [phenol sulfuric acid (PSA) for polysaccharides; Coomassie blue method for proteins].
• Fluorescence microscopy
• Atomic Force Microscopy (AFM)
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EIS in-situ (day 1)
31
Nyquist
Bode
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EIS in-situ (day 3)
32
Bode
Nyquist
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EIS in-situ (day 7)
33
Nyquist Bode
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EIS modeling for in-situ
34
RS(QP(RP(QCTRCT))) model:
RS = solution resistance
QCT = charge transfer CPE
RCT = charge transfer resistance
QP = biofilm pores CPE
RP = biofilm pore resistance
CPE = constant phase element
RS (ohm) CPEP n (0<n<1) RP CPECT NCT
0<n<1
RCT
PAO1 day 1 9.94 6.999×10-4 0.8926 8.92 6.716×10-4 0.8797 18020
PAO1 day 3 10.18 6.432×10-4 0.9173 10.21 6.934×10-4 0.9143 31560
PAO1 day 7 10.44 1.833×10-4 0.9532 12.48 6.967×10-4 0.8078 80560
SRB day 1 5.49 4.345×10-4 0.9878 4.64 8.125×10-4 0.8189 6952
SRB day 3 6.84 1.236×10-4 0.9938 3.24 3.048×10-4 0.9972 6592
SRB day 7 6.11 1.551×10-4 0.8597 1.03 9.493×10-5 0.8696 3603
Mixed day 1 8.38 9.061×10-4 0.8362 5.24 3.899×10-4 0.8018 12130
Mixed day 3 9.24 1.585×10-4 0.8231 8.15 4.334×10-4 0.8213 17680
Mixed day 7 8.73 4.933×10-4 0.8723 6.92 4.231×10-4 0.8419 15020
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Interfacial Resistances (RCT and RP) : in-situ
35
RP RCT
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Potentiodynamic polarization : in-situ
36
Ecorr (mV) Icorr (µA/cm2) βa βc
PAO1 -812 0.118 43.2 × 10-3 82 × 10-4
SRB -894 0.761 68.4 × 10-3 46.1 × 10-3
Mixed -845 0.424 73 × 10-4 52 × 10-4
The reduction in RCT and Icorr for mixed bacteria
cultures compared to SRB can be attributed to
(i) Electron transfer competitions between NO3- & SO4
2- in the presence of PAO1
(ii) Inhibitory effects of denitrification pathway products (ex. Nitrite) for SRB
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EIS ex-situ (day 1)
37
Nyquist Bode
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EIS ex-situ (day 3)
38
Bode Nyquist
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EIS ex-situ day (7)
39
Nyquist Bode
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EIS ex-situ modeling
40
RS(QP(RP(QCTRCT))) model:
RS = solution resistance
QCT = charge transfer CPE
RCT = charge transfer resistance
QP = biofilm pores CPE
RP = biofilm pore resistance
CPE = constant phase element
RS CPEP n 0<n<1 RP CPECT nCT 0<n<1 RCT
PAO1 day1 1.524 1.79×10-7 0.9436 1.582 9×10-3 0.9328 60.63
PAO1 day 3 1.237 2.18×10-7 0.9081 1.327 1.4×10-2 0.907 88.13
PAO1 day 7 1.069 9×10-7 0.9274 1.365 3.46×10-2 0.9992 88.94
SRB day 1 0.8252 9.7×10-8 0.8694 42.56 8.4×10-4 0.9931 234.6
SRB day 3 0.8502 1.1×10-7 0.8607 18.2 1.3×10-2 0.8571 197.2
SRB day 7 0.9869 8.7×10-8 0.8932 63.1 9.3×10-3 0.8777 247.1
Mixed day 1 0.7114 9.7×10-8 0.8432 4.45 5.4×10-3 0.87 207.4
Mixed day 3 2.746 1.65×10-7 0.9438 5.261 7.2×10-3 0.9962 65.23
Mixed day 7 0.9632 7.8×10-8 0.931 7.611 5.6×10-3 0.917 152.8
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Interfacial Resistances (RCT and RP) : ex-situ
41
RP RCT
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Potentiodynamic polarization : ex-situ
42
Ecorr (mV) Icorr (µA/cm2) βa βc
PAO1 -814 9.43 87.6×10-3 33.35×10-2
SRB -730 3.77 48.4×10-3 57.5×10-3
Mixed -773 7.82 53.6×10-3 73.8×10-3
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EIS ex-situ sterile
43
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EIS ex-situ sterile modeling
44
RS(QCTRCT) model:
RS = solution resistance
QCT = charge transfer CPE
RCT = charge transfer resistance
CPE = constant phase element
Days RS CPE n (0<n<1) RCT
Day 1 0.86 2.94×10-3 0.9629 278.5
Day 3 0.89 1.62×10-3 0.9267 193.6
Day 7 0.81 3.78×10-3 0.9639 174
SRB biofilm protects the surface since RCT,SRB>RCT,sterile
PAO1 biofilm cannot protect the surface R
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Fluorescence microscopy and EPS
quantification 45
0
10
20
30
40
50
60
70
80
90
SRB Mixed PAO1
% Area Polysaccharide
Protein
Total coverage
SRB
PAO1
Mixed
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Fluorescence microscopy and EPS
quantification, cont’d
46
Cultures
Polysaccharide
(µg / cm2 coupon)
Protein
(µg / cm2 coupon)
PAO1 23.4 ± 2.8 7.6 ± 1.3
SRB 59.2 ± 2.9 86.8 ± 18.1
Mixed 47.7 ± 7.7 76.22 ± 34.3
Protein and polysaccharide content as well as
Surface coverage is higher for SRB compared to
mixed species and PAO1 biofilms respectively
At high iron concentrations on the surface of
coupons PAO1 attachment to iron surface decreases
resulting in patchy and scanty biofilms
0
10
20
30
40
50
60
70
80
90
SRB Mixed PAO1
% Area Polysaccharide
Protein
Total coverage
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Atomic Force Microscopy (AFM)
AFM was used to:
(i) Compare the influence of bacterial metabolites in PAO1 and SRB solutions on
corrosion behavior of cast iron (in-situ)
(ii) Study effect of biofilms on corrosion behavior of cast iron in a standard chloride
solution (3.5% NaCl solution, pH = 4, ex-situ).
7 days of biofilm development followed by biofilm removal by Clark’s solution.
47
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Atomic Force Microscopy (AFM) :
metabolites effect, in-situ
48
PAO1;
Roughness
SRB;
Roughness
Pits
Ra = 14.94 nm
Rq = 23.45 nm
Ra = 38.36 nm
Rq = 66.24 nm
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Atomic Force Microscopy (AFM) :biofilm
effect, ex-situ
49
PAO1;
Roughness
SRB;
Roughness Ra = 664.9 nm
Rq = 845.4 nm
Ra = 110.5 nm
Rq = 146.3 nm
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Atomic Force Microscopy (AFM)
High surface roughness and distorted surface morphology after PAO1 biofilm removal in
coupons that have been immersed for further 4 days in 3.5% NaCl solution compared to
SRB biofilms. This reveals weaker ability of PAO1 biofilms compared to SRB in
protecting the surface.
Pits and higher roughness for coupons that have been immersed in SRB solutions for 7
days without immersion in NaCl. This reveals detrimental effect of SRB metabolites (such
as H2S) in its bacterial solutions compared to PAO1.
50
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Weight loss
Weight loss was used to grasp an idea about the actual extent of material loss due to
corrosion. ASTM D-2688.
It was conducted to study effect of bacterial metabolites in solution and biofilms alone.
7 days of biofilm development on coupons followed by biofilm removal:
(a) without immersion in 3.5% NaCl solution Influence of bacterial metabolites.
(b) immersion in 3.5% NaCl solution for 4 days Effect of biofilm.
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Weight loss
52
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
Day 1 Day 3 Day 7
Weight loss
mg/cm2
SRB
PAO1
Mixed
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
Day 1 Day 3 Day 7
Weight loss
(mg/cm2)
SRB
PAO1
Mixed
Bacterial metabolites effect (in-situ) Biofilm effect (ex-situ)
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Outline
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Introduction and Theoretical background
Scope of Study Materials and
Methods Results and Discussions
Conclusions
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Conclusions
Mixing denitrifying PAO1 with SRB lead to a decrease in the extent of corrosion of cast
iron (higher electrical resistances, less weight loss) compared to SRB monocultures which
can be attributed to reduced SRB activity in the presence of PAO1.
Although SRB bacterial solution exerted higher corrosion on coupons, SRB biofilms
protect the surface because of their high surface coverage and weight in the surface of cast
iron.
Surface roughness and pitting is higher in SRB solutions mostly because of H2S, however
SRB biofilms could protect the surface against attack of an aggressive electrolyte.
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Acknowledgements
I appreciate:
Dr Youngwoo Seo (My Adviser)
This research was conducted under the support of NSF award # 1236433.
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