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Contributions of CINVESTAV to the study of bioreactors with
simultaneous electron acceptors
Removal of Trichlorophenol under partially-aerated methanogenic using a Fluidized Bed bioreactor
Removal of Perchloroethylene in partially-aerated methanogenic regime using a Fluidized Bed bioreactor
Removal of Perchloroethylene under M-D conditions
M-A and M-D bioreactors coupled to zero valent iron for PCE degradation
Removal of perchloroethylene in two methanogenic-denitrifying
continuous systems
Héctor M. Poggi-Varaldo
CINVESTAV-IPN, Dept. Biotechnology and Bioengineering, Environmental Biotechnology R&D Group, México
Contents
Acknowledgements Abbreviations Introduction
– Perchloroethylene– Simultaneous Electron Acceptor (SEA)
systems Methodology
– Reactors set-up and operation– Methods
Results and discussion Conclusions
Acknowledgements
Orgazing Committee and ENCB-IPN CINVESTAV, CONACYT Mr. Claudio Garibay-Orijel,
Mr. Rafael Hernández-Vera, Ms Paola Zárate-Segura
Prof. Elvira Ríos-Leal Dr. Jaime García-Mena
Abbreviations
Bv organic loading rate per unit volume
CM complete mix reactor
DCE dichloroethylene
FBBR anaerobic fluidized bed biological reactor
Ig biogas productivity
PCE perchloroethylene
TCE trichloroethylene
VC vinyl chloride
alpha factor
net increase
removal efficiency
loading ratio of organic matter as COD to nitrogen-nitrate
v PCE loading rate per unit volume
Introduction
Perchloroethylene
•potentially hazardous
•included in the priority list of hazardous pollutatns of USA and other countries (EPA, 1993)
•widely used as a solvent in dry-cleaning industry as and a degreasing solvent in the metal and machinery industries for more than 50 years (HSIA, 1999)
•in the 90’s, a world demand between 160 000 and 520 000 tonnes/yr has been recorded (EPA, 1993; WHO, 2000).
•PCE is recalcitranr (not biodegradable) in aerobic conditions (Vogel and McCarty 1985). Its biological transformation is generally carried out in anaerobic environments (van Eekert, 1999).
•Previous works on biological treament and bioremediation of PCE have used anaerobic consortia that mediated the sequential reductive dehalogenation of PCE, presumably by cometabolism (Vogel y McCarty, 1985; Prakash y Gupta, 2000; Cope et al., 2001; López-Navarrete et al.,
2003).
•In most of these studies, accumulation of dichloroethylene (DCE) and vinyl chloride (VC) has been observed. A few works have been able to show the full transformation of PCE to ethene, after a long enrichment of anoxic consortia or using pure cultures of dehalorespiring bacteria (Mayor et al., 2002).
Series reactorsand
Simultaneous Electron Acceptor (SEA) systems
High chlorine content
Penta-, tetra-, tri-
Anaerobic
Low chlorine content
di-, and monochloro-
AerobicAccumulatio
n
of
A good alternative: Series Reactors
Brief example with a chlorinated organic compound
Series Reactors
Anaerobic Reactor
Reactor with second electron acceptor
Tetrechloroethene
TCE, DCE, VC
Further removal of chlorinated aliphatics
Garibay Orijel et al. (2005a) J. Chem. Technol. Biotechnol. In pressCampos-Velarde et al. (1997). Battelle Press.
What’s the problem with series reactors?
2 Reactors
Costs X 2
Better try Simultaneous
Electron Acceptors in One reactor
Salto a la albóndiga
Protection via diffusion barrierin biofilm
CH4
CO2
NO3- + org. matter
N2 +H2O
An
aero
bic
zo
ne
Den
itri
yin
g
zon
e
Liq
ue
d d
if-
fusi
on
lea
yer
Denitrifying microorganisms
Methanogens
Bu
lk l
iqu
id(adapted from López-Navarrete, 2002).
Pollutant
NO3-
Concentration
Carrier
= g COD/g 2nd electron acceptor in the influent
2nd electron acceptor: NO3-
• lambda determines the percentage P of substrate that is channelled into methanogenesis or denitrification in SEA conditions
• it can be demonstrated that P = 50% at = 9 for M-D reactors
Garibay-Orijel et al. (2005b). J. Chem. Technol. Biotechnol.
Objectives
•to evaluate and compare de performance of a fluidized bed bioreactor (FBBR) and complete mix reactors (CM) with suspended biomass, all of them fed with PCE as model chlorinated aliphatic and a low-moderate concentration of degradable organic matter as methanol
• to assess the influence of the biochemical regime (full methanogenic versus M-D), and the effect of in the M-D regime (18 and 9) on performance
Methodology
Nmin
1
2
3
5
7
8
9
6
4
2´
A
Nmin
1
2
3
5
6
7
4
2´
N. max
B Reactor setup: A Anaerobic Fluidized Bed Bioreactor (AFBBR); B Complete Mix Bioreactor (CM). 1A fluidized bed of bioparticles; 2A reservoir and feed of influent with a partial content of methanol; 2´A feed of stock of PCE in methanol; 3A recirculation; 4A liquid trap; 5A effluent reservoir; 6A biogas exit; 7A biogas sampling port; 8A activated carbon trap; 9A biogas measurement by brine meters; 1B suspended biomass; 2B reservoir and feed of influent with a partial content of methanol; 2´B feed of stock of PCE in methanol; 3B effluent reservoir; 4B biogas exit; 5B biogas sampling port; 6B activated carbon trap; 7B biogas sampling port.
Anaerobic fluidized bed reactora Complete Mix Reactorb Complete Mix Reactor2c
Bvd
(gCOD/Ld)
ve
(mg PCE/Ld)
PCEf
(mg/L)
(gCOD/
gN-NO3)
Bvd
(gCOD/Ld)
ve
(mg PCE/Ld)
PCEf
(mg/L)
(gCOD/
gN-NO3)
Bvd
(gCOD/Ld)
ve
(mg PCE/Ld)
PCEf
(mg/L)
g (gCOD/
gN-NO3)
Period 1 1.0 40 40 0 0.066 1.33 20 0 0.066 2.66 0 0
Period 2
1.0 40 40 18 0.066 1.33 20 18 0.066 2.66 40 18
Period 3 1.0 40 40 9 0.066 1.33 20 9 0.066 2.66 40 9
Notes.
1000 mg COD-methanol/L in all periods; PCE: Perchloroethylene.a Anaerobic fluidized bed bioreactor at HRT= 1d, Vop= 2.8L, 35°C; b,c Complete mix reactor at HRT= 15 d, Vop= 2.5L, 35°C.; d Volumetric loading rate of organic matter in gCOD/(L.d), e Volumetric loading rate of PCE in mg PCE/(L.d); f Concentration of PCE in the influent, in mg/L; g Relation of volumetric loading and Nitrogen contend in Nitrate.
Operating conditions of bioreactors in the different periods of the experimental design
Period 1
Period 2
Period 3
40
Operation and monitoring
mesophilic conditions glass column (2.8 L), loaded with 1 L of 1 mm
granular activated carbon, colonized by an anaerobic consortium
methanol (1000mgCOD /L) hydraulic residence time = 1 d (bed basis)
Response variables:– Removal efficiency of organic matter (COD)– Removal efficiency of PCE– Concentration of less substituted chlorinated aliphatics– Specific methanogenic activity, specific denitrifying
activity, specific oxgyen uptake rate
T: transient period with 2vvd
HRT= 1 d, TCPi = 80mg/L, Phei = 20mg/L, CODi =1000 mg/L, mesophilic
15vvd
1 2 3
50
75
100
90 140 190 240 290
D
QO
( %
)
0
10
20
90 140 190 240 290
C
l- (m
g/L
)
0
0.2
0.4
90 140 190 240 290
Ig (L
bio
gas/ L
reacto
r d
)
A
B
C
Dynamic performance of reactors
A: Removal efficiency of COD versus time;
B Biogas productivity;
C Increase of chloride anion.
FBBR
○ CM 1
∆ CM 2
M = 18 = 9
Fluidized bed reactor
Fluidized bed reactor
Complete mix reactors
Complete mix reactors
Complete mix reactors
CO
D (
%)
I g (
L b
iog
as/L
.d)
∆C
l- (m
g/L
)
Time (day)
Average performance of reactors 1/2 Period Parameter 1 2 3 Reactor
PCE (%) a 98.81 1.15 98.67 2.22b
98.62 2.10
98.88 1.12 98.64 3.89 98.10 2.81
98.60 5.95 92.39 2.80 90.01 ( 1.13
AFBBR c CM 1 d CM 2 e
COD (% ) f 94.66 5.21 92.49 2.10b 73.46 2.15
98.85 6.40 96.21 3.33 91.73 3.80
98.87 1.51 96.61 7.35 97.30 8.53
AFBBR CM 1 CM 2
NO3 (% ) g ---- ---- ----
97.88 2.56 99.58 4.34 98.71 3.82
98.91 0.27 98.43 1.31 97.79 0.87
AFBBR CM 1 CM 2
ΔCl- (mg/L) h 11.7 5.3 9.9 3.2b 7.5 5.2
14.53 9.4 9.78 4.2 9.99 3.3
18.7 0.7 16.21 0.8 17.3 0.6
AFBBR
CM 1
CM 2 i 0.30 0.01
0.39 0.01b
0.30 0.01
0.24 0.018 0.28 0.02 0.30 0.02
0.25 0.05 0.28 0.04
0.27 0.05
AFBBR
CM 1
CM 2 CH4 (%) 70.20 7.32
40.30 1.03b
30.37 1.23
60.52 3.65 48.5 4.34 49.5 4.78
65.4 4.45 10.10 3.21
8.50 3.25
AFBBR
CM 1
CM 2 Ig(L/L.d) j 0.41 0.015
0.11 0.01b
0.0140.03
0.39 0.01 0.12 0.01 0.04 0.03
0.280.02 0.022 0.02
0.026 0.02
AFBBR
CM 1
CM 2 VSS (mg/L)k 58.34 2.40
240 3.45b 278 3.65
56.66 ± 3.43 430 5.34 475 5.16
71.34 3.45 420 5.35 410 5.21
AFBBR CM 1 CM 2
NKTbp (mgN/gbpseca)l NKT vss (mgN/L) NKT vss (mgN/L)
11.26 ( 1.23 30.08 ( 0.67b 34.75 ( 0.65
25.45( 2.32 38.72( 3.24 37.23 5.13
32.78 3.24 43.74 2.35 42.45 4.23
AFBBR
CM 1
CM 2 x (mg PCE/g VSS d)m x (mg PCE/g VSS d) x (mg PCE/g VSS d)
17.2 1.2 5.5 0.7 9.6 0.7
7.64 1.3 4.28 0.9 8.91 0.4
5.93 0.3 3.16 0.5 7.81 0.3
AFBBR
CM 1
CM 2 SMAn (mmol CH4/g vss h)
0.060 0.02 0.067 0.03 0.058 0.04
0.067 0.03 0.065 0.03 0.033 0.01
0.075 0.043 0.010 0.004 0.022 0.012
AFBBR
CM 1
CM 2 SDAo (gNO3
- /gNKT h) 0.17 0.09 0.27 0.12 0.20 0.10
0.17 0.05 0.25 0.07 0.20 0.02
0.29 0.02 0.17 0.04 0.15 0.08
AFBBR
CM 1
CM 2
Average performance of reactors 2/2Metabolites and dechlorination efficiencies
Period
Parameter
1
2
3
Reactor
Metabolites in the effluent PCE ( mol/L) a 2.85 4.1
3.2 2.9 3.3 0.7
3.14 2.1 3.3 1.2 4.6 2.1
3.3 1.2 18.4 2.1 23.6 2.4
AFBBR c CM 1 d CM 2 e
TCE ( mol/L) f 62.7 8.2 29.6 16.0b 134.8 12.8
12.3 1.3 22.91 12.2 49.12 5.4
11.5 3.2 10.8 2.1 30.4 5.9
AFBBR c
CM 1 d
CM 2 e
DCE (mol/L) g 4.1 1.5 2.6 1.2b
0.5 0.4
undetectable 8.99 4.5
41.92 2.0
18.1 2.8 25.1 1.4 35.0 3.5
AFBBR c
CM 1 d
CM 2 e
VC (mol/L) h 127.6 4.3 138.5 8.5b
85.7 0.6
125.9 9.7 138.7 3.2
154.0 10.9
74.6 10.1 79.58 9.2 160.4 9.4
AFBBR c CM 1 d CM 2 e
Metabolites by stripping PCE (mol/gAC) 1.1E-2 1.1E-2
1.7E-4 5.0E-4b
9.1E-4 7.2E-4
1.1E-2 7.4E-3 1.7E-3 1.0E-4 9.0E-4 1.1E-4
1.1E-3 1.2E-2 1.7E-3 1.4E-3 9.0E-4 4.5E-4
AFBBR c
CM 1 d
CM 2 e
TCE (mol/ gAC ) 3.6E-2 6.3E-3 1.6E-4 1.2E-4b 3.1E-4 1.1E-4
2.7E-2 1.4E-2 1.0E-4 0.5E-4 2.3E-4 1.3E-4
1.2E-2 0.4E-2 1.1E-4 0.3E-4 2.3E-4 1.3E-4
AFBBR c
CM 1 d
CM 2 e
DCE (mol/ gAC ) 3.5E-2 3.2E-3 undetectableb
1.0E-3 1.2E-3
1.7E-2 1.1E-2 undetectable
5.12E-4 3.2E-4
3.2E-3 2.5E-3 undetectable
5.1E-3 3.2E-3
AFBBR c
CM 1 d
CM 2 e
VC (mol/ gAC) 1.1E-1 1.2E-2 2.7E-4 1.4E-4b
5.1E-2 1.2E-4
2.8E-2 1.3E-2 4.2E-3 2.1E-3 1.3E-2 0.9E-2
6.3E-3 2.1E-3 4.1E-3 2.3E-3 1.2E-2 1.0E-2
AFBBR c
CM 1 d
CM 2 e
Adsorption of metabolites on the bioparticles
PCE (mol/ gbp) 1.7E-1 2.1E-2 1.8E-1 3.03E-2 4.4E-1 1.63E-3
TCE (mol/ gbp) 1.4E-3 0.3E-3 undetectable 5.7E-1 1.63E-3
DCE (mol/ gbp) 7.1E-1 1.1E-2 3.6E-2 1.15E-2 5.5E-1 3.19E-2
VC (mol/ gbp)
7.3E-1 6.2E-2 9.6E-1 1.42E-1 1.2 1.42E-1
AFBBR e
Declorination Efficiency CL b(%) j 64.19 2.19
48.94 1.98 47.49 4.21
81.86 3.21 50.33 5.54 57.78 3.53
83.39 3.21 57.56 2.43 56.63 4.31
AFBBR c
CM 1 d
CM 2 e
Poggi’s discrete divergence index
n’A = only green
n’B = only yellow
nA = green plus white
nB = yellow plus white
Microbial community
A
Microbial community
B
Zárate-Segura et al. (2005). Battelle
Poggi’s discrete divergence indexand dynamic divergence coefficient
Poggi = (n’A + n’B)/(nA + nB)
wherewhere
n’n’AA = number of bands in A that are not in B = number of bands in A that are not in B
n’n’BB = number of bands in B that are not in A = number of bands in B that are not in A
nnAA = total number of bands in A = total number of bands in A
nnBB = total number of bands in B = total number of bands in B
complete similarity 0 complete similarity 0 PoggiPoggi 1 complete divergence 1 complete divergence
Poggi = d(Poggi)/dt; dynamic divergence coefficient
.
Zárate-Segura et al. (2005). Battelle
Lanes 1 and 2, methanogenic period with no PCE; Lanes 3 to 5, methanogenic period with 20 mg/L PCE; Lanes 6 to 8 methanogenic period with 40 mg/L PCE. All lanes contain 5 g of 16S rDNA PCR product. Poliacrylamide gel at 8% (Acrylamide/N-N´methylenbisacrylamide 37.5:1) Buffer TBE 1X, Urea -Formamide 10-50% (8M of Urea and 40%v/v formamide equivalent to 100% denaturing agents); 30 V, 13-15 mA, 8 h, 60°C.
DGGE profiles of major bacterial communities present in fluidized bed bioreactor
∆Poggi Jaccard index of similarity
Index Reactor FBBRa CM1a CM2a FBBRa CM1a CM2a FBBR 0.05
0.71
0.44
0.73
0.17
0.39
CM1 ------
0.33
0.67
------
0.50
0.20
CM2 ------ ------ 0.67 ------
------
0.20
Variation of bacterial communities in methanogenic FBBR and CM reactors in a period of operation with no PCE in the influent:
Diagonal: comparison between start and end of the period for a given bioreactor
Other cells: comparison between bioreactors
∆Poggi Jaccard index of
similarity Index Reactor FBBRa CM1a CM2a FBBRa CM1a CM2a FBBR 0.05 0.65
0.65
0.91
0.22
0.22
CM1 ------
0.00
0.86
------
1.00
0.08
CM2 ------ ------ 0.50 ------
------
0.32
Variation of bacterial communities in methanogenic FBBR and CM reactors in a period of operation with 40 mg PCE/L for
FBBR and CM2 and 20 mg PCE/L in CM1:Diagonal: comparison between start and end of the period for a given bioreactor Other cells: comparison between bioreactors
∆Poggi Jaccard index of
similarity Index Reactor FBBRa CM1a CM2a FBBRa CM1a CM2a FBBR 0.10
------
------
0.82
------
------
CM1 ------
0.63
------
------
0.22
------
CM2 ------ ------ 0.60 ------
------
0.25
Variation of bacterial communities in methanogenic FBBR and CM reactors between periods with no PCE and with PCE
in the feed:Diagonal: comparison between periods for a given bioreactor Other cells: empty
System Poggi (1/day) Reference
Methanogenic fluidized bed bio-reactor, 1000 mg/L COD-MeOH
0.0003 This work
Methanogenic complete mix bio-reactor, 1000 mg/L COD-MeOH
0.0022 This work
Methanogenic complete mix bio-reactor, 1000 mg/L COD-MeOH
0.0045 This work
Sequencing batch reactor treating piggery wastewater
0.260 1
Membrane-coupled bioreactor dedi-cated to biological nutrient removal
0.084 2
UASB reactor treating a penta-chlorophenol-contaminated wastewater
0.010 3
. .
Ref.: 1. Juteau, P., Tremblay, D., Villemur, R., Bisaillon, J.-G. and Beaudet R. (2004). Analysis of the bacterial community inhabiting an aerobic thermophilic sequencing batch reactor (AT-SBR) treating swine waste. Appl. Microbiol. Biotechnol. 66, 115-122.
2. Ghosh, S. and LaPara, T.M. (2004). Removal of carbonaceous and nitrogenous pollutants from a synthetic wastewater using a membrane-coupled bioreactor. J. Industrial Microbiol. Biotechnol. 31, 353 –361.
3. Tartakovsky, B., Manuel, M.F., Beaumier, D., Greer, C.W. and Guiot, S.R. (2001). Enhanced selection of an anaerobic pentachlorophenol-degrading consortium. Biotechnol. Bioeng. 73, 476-483.
Dynamic divergence indices of bacterial communities of several bioreactors
Conclusions
- In Period 1, methanogenic regime, AFBBR showed the best performance of the three reactors with higher values of both organic matter removal and PCE, in spite that volumetric loadings on AFBBR were 15-fold higher and some process stress would have been expected.
- During Period 2, simultaneous M-D regime at =18 gCOD/gN-NO3
-, an improvement in performance of CM2 was observed. The other two reactors displayed similar performances than the corresponding in Period 1.
-In Period 3, simultaneous M-D regime at = 9 gCOD/gN-NO3
-, and concerning the two reactors fed with the highest PCE concentration 40 mg/L (FBBR and CM2), the FBBR outperformed CM2 in almost all the performance parameters and its metabolite profile was better than both of CM2 and CM1 (lower TCE, DCE, and VC in the effluent).
This pattern, along with highest dechlorination efficiency of FBBR strongly suggests that FBBR under M-D conditions may provide a more integral treatment to wastewaters polluted with significant concentrations of PCE.
– The bacterial communities in the CMs were richer than those of FBBR during three operations periods
– Generally, bacterial profiles in each reactor varied with time in a given period of operation. There was a relative higher stability of consortium in the FBBR as displayed by the lowest dynamic divergence coefficients values
– The above described pattern was accompanied by a better biochemical peformance of FBBR (stable methanogenesis, high removal of PCE and lower concentrations of intermediate chlorinated aliphatics)
– Despite relative variation of bacterial consortia with time, bioreactors showed steady state biochemical performance
– PCE had a negative impact on the richness of CMs consortia whereas this impact was less noticeable in the FBBR.
Questions and alibishectorpoggi2001@yahoo.com
Despedida Farewell
Me miro al espejo I look at myself in the mirrory sólo veo but I can only seela desnuda pared a mis espaldas. the bare wall behind me.Me estremezco y me pregunto: I shiver and mutter:¿quién soñará nuestros sueños? Who will dream our dreams?¿quién peleará nuestras batallas? Who will fight our battles?¿quién llorará nuestras derrotas? Who will cry for our defeats?¿quién ganará nuestras victorias? Who will win our victories?
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