rhizosphere-enhanced treatment of pahs at cold, · pdf filerhizosphere-enhanced treatment of...
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![Page 1: Rhizosphere-Enhanced Treatment of PAHs at Cold, · PDF fileRhizosphere-Enhanced Treatment of PAHs at Cold, Remote Locations; Challenges of Application and Monitoring Mike Reynolds,](https://reader031.vdocuments.mx/reader031/viewer/2022011723/5a9c9cfe7f8b9a01398b7280/html5/thumbnails/1.jpg)
Rhizosphere-Enhanced Treatment of PAHs at Cold, Remote Locations;
Challenges of Application and Monitoring
Mike Reynolds, Larry Perry, Karen Foley, Dave Ringelberg, Brent Koenen, and Kevin McCarthy
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Federal Cold Region Sites Low Input Systems
Low Input SystemsHigh Input Systems
Natural attenuation
IncinerationDig and haul
LandfarmingAir sparging and bioventing
CompostingBioslurry reactors
Low-temperature thermal desorptionSoil washing
COST
Phytoremediation / Rhizosphere enhancement
T I M E
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Ground Water
Saturated Zone
Relatively constant conditions
Source Contamination
~~ Mixing ! Natural Attenuation
Water Soluble Contaminants
Relatively constant temperature and moisture
Surface soil is less physically mixed than GW, and undergoes temperature, moisture, carbon and nutrient changes
PermafrostActive Zone
Wide-scale, low-cost, in-situ treatment� Requires less input� Requires fewer specifics about a
site
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Problem� Remote locations� Hundreds of sites� Cleanup is costly
� Mobilization & demobilization� Limited alternatives� Short season� Relatively slow activity
Problem� Remote locations� Hundreds of sites� Cleanup is costly
� Mobilization & demobilization� Limited alternatives� Short season� Relatively slow activity
Limitations� Temperature� E-S proximity, MT limitations� Microorganisms (numbers,
diversity, activity, function)� Nutrient & contaminant
availability, bioavailable C� Aeration
Limitations� Temperature� E-S proximity, MT limitations� Microorganisms (numbers,
diversity, activity, function)� Nutrient & contaminant
availability, bioavailable C� Aeration
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Time
Rel
ativ
e C
once
ntra
tion
of C
onta
min
ant (
% o
f Co)
0
20
40
60
80
100
Typical Bioremediation
Ideal Bioremediation
Impact of Limitations
Lag may be longer
Residual may be higher
Slope may be shallower
Microbial activity in soil is not constant �but starts and stops many timesf (temperature, moisture, carbon additions, ???)
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� Analog enrichment (natural forced molecular evolution)
� Stimulated microbial #s & activity
� Reduced M-T limitations, pseudo-mixing
� Carbon-enriched environment
� Not necessarily plant uptake
� Increased OM - Humification
� ???
Stimulating Microbial Activity via the
Rhizosphere Effect
CO2CO2
CO2CO2
CO2
Rhizosphere Fallow
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Variability in Contaminant and Productconcentrations makes their routine use for monitoring difficult, and this is exacerbated for:
� Surface soils� Non �brute-force� treatment
methods� Most situations � whether
implementation, regulatory monitoring, or research studies
Reynolds, C. M. 1993. Field measured bioremediation rates in a cold region landfarm: Spatial variability relationships.
Landfarm at Fairbanks Airport� Tilled ~ weekly� Fertilized and limed� Irrigated� Periodically, composite samples
taken near 25 nodes� Calculated half-lives varied ~7-X
Measuring Contaminant Loss is Difficult
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Fairbanks Site
ReplicatedGrasses, Nutrients, Both, Control�Soil sock� approachCrude and Diesel
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Vegetated
The rhizosphere�effect really does work � Fairbanks data
ti
tf
Control
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\HPC
HEM
\...\046R0101.D
Time (min.)
10 20 30 40
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\064R
0101.D
Time (min.)
10 20 30 40
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\067R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\070R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\077R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4Sig. 2 in E:\FID
54\080R0101.D
Time (min.)
T=1
Fall 1995
T=2
Spring 1996
T=3
Fall 1996
T=4Spring 1997
T=5Fall 1997
T=0
Summer 1995
T=5
Fall 1997
10 20 30 40
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\064R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\085R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\088R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\095R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\098R
0101.D
Time (min.)
10 20 30 400
10000
20000
30000
40000
Sig. 2 in C:\H
PCH
EM\...\052R
0101.D
Time (min.)
T=5
Fall 1997
T=1
Fall 1995
T=2
Spring 1996
T=3
Fall 1996
T=0
Summer 1995
T=4
Spring 1997
~700 days
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TPH Changes
Days0 28 56 84 112 140 168 196
TPH
(mg/
kg)
0
1000
2000
3000
4000
5000
6000
Vegetated
SterileUnvegetated
Lab studyAlaska soilWinter ryeNo nutrients added
Significant plant effect
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Hexadecane
Days0 28 56 84 112 140 168 196
0
200
400
600
800
Sterile
Unvegetated
Vegetated
Easy to degrade,Little difference
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Pyrene
Days0 28 56 84 112 140 168 196
Pyre
ne (m
g/kg
)
0
200
400
600
800
Vegetated
UnvegetatedSterile
Recalcitrant, big difference
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Selected CompoundsNon-vegetated
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Selected CompoundsVegetated
Selected CompoundsNon-vegetated
We can subtract Vegetated from Non-Vegetated to visual rhizosphere effect
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Benefit due to Rhizosphere Effect [ Non-veg ] - [ Vegetated ] is f (Compound and Time)
Rhizosphere effect has Time and Compound Component s
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Petroleum - a Complex Mix of hydrocarbons
AsphaltenesPolars
AromaticsPAH
Saturates
Volatiles
Compositional Variability� May consist of 100s to 1000s of
hydrocarbons� Natural or remediation-enhanced weathering
of hydrocarbons in soil substantially alters its composition
� Chromatographic methods yield most accurate measures of extractable hydrocarbons
� Inherent variability in TPH can be minimized by normalization to a recalcitrant marker compound
Generalized Composition of Crude Oil
Biomarkers�
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Benefits to Normalizing TPH to Hopane
� Precision (concentration data) can be increased when data are normalized to internal marker such as hopane
� Raw TPH soil data has inherently high variability
� Concentration data can become more variable as petroleum weathers
� Normalizes TPH concentration data
0102030405060708090
100
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19
Months
Depl
etio
n of
Tot
al H
ydro
carb
ons
Raw TPH data
NormalizedTPH data
Assumes that contaminant at a site has uniform composition at t=0
Biomarkers�
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In addition to TPH, we can normalize other compounds w.r.t a biomarker
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Two Field Demo Sites in Korea
Not cold sites, but in practice similar problems
to cold regions
FactorialRyegrass (Yes / No)Nutrients (Yes / No
4 repsComposite samples
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Monitoring Rhizosphere-enhanced remediation�
� Petroleum� 1000s of compounds� biodegrade at different rates� by different enzymes
� We can exploit differential rates� confirm degradation� compare treatments
� And it may also give us better insight
C P F P+F
KoreaSite 2
� These data further demonstrate rhizosphere-enhancement in field
Benefit due to Rhizosphere Effectis f (Compound and Time)
Lab Studies
Time
Recalc
itrance
�Response� or Control -
Treatment
$ Data also suggest that for effective monitoring, you must know:� How to sample� What to look for � When to look for it�
� and when is f (microbial processes), not our calendars
� � and microbial processes are f (conditions)�{H2O, Temp., Carbon additions}
How can we do this?
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Campion
FactorialVegetation (Yes / No)Nutrients (Yes / No
4 repsComposite and soil sock samples
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Campion
� �Bottom� of bioventing pile� Previously fertilized and treated
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-350%-300%-250%-200%-150%-100%
-50%0%
50%100%150%200%
C F F+P P
Campion, TPH DepletionsCampion � TPH Depletions
Campion, t1-t2, PAH Depletions
-600%-500%-400%-300%-200%-100%
0%100%200%300%
C F F+P P
Campion � PAH Depletions
� Weathered at t=0
� �Polishing�� Most effect is
with PAHs
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Annette Islandactorial
Vegetation (Yes / No)Nutrients (Yes / No
repsomposite and soil sock samples
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Annette Island
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Annette TPH Depletions
-350%-300%-250%-200%-150%-100%
-50%0%
50%100%150%
C F F+P P
Annette Island � TPH Depletions
C PF+PF
Annette % PAH depletion t1 to t4by treatment
-400%
-300%
-200%
-100%
0%
100%
200%
300%
C F F+P P
Annette Island � PAH Depletions
� Non-uniform composition
� Nutrients and (Nutrients and Plants) starting to respond
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Barrow
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Barrow
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Barrow, t1-t4, %TPH Depletion
-80%-60%-40%-20%
0%20%40%60%
C F PF+P
Barrow - TPH Depletions
Barrow, t1- t4, %PAH Depletion
-20%
0%
20%
40%
60%
80%
C F F+P P
Barrow � PAH Depletions
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Non-Freezing Degree Days
010002000300040005000600070008000
Campion Annette Island Barrow
-350%-300%-250%-200%-150%-100%
-50%0%
50%100%150%200%
C F F+P P
Campion, TPH DepletionsCampion � TPH Depletions
-350%-300%-250%-200%-150%-100%
-50%0%
50%100%150%200%
C F F+P P
Campion, TPH DepletionsCampion � TPH Depletions
Campion, t1-t2, PAH Depletions
-600%-500%-400%-300%-200%-100%
0%100%200%300%
C F F+P P
Campion � PAH DepletionsCampion, t1-t2, PAH Depletions
-600%-500%-400%-300%-200%-100%
0%100%200%300%
C F F+P P
Campion � PAH Depletions
Annette TPH Depletions
-350%-300%-250%-200%-150%-100%
-50%0%
50%100%150%
C F F+P P
Annette Island � TPH DepletionsAnnette TPH Depletions
-350%-300%-250%-200%-150%-100%
-50%0%
50%100%150%
C F F+P P
Annette Island � TPH Depletions
Annette % PAH depletion t1 to t4by treatment
-400%
-300%
-200%
-100%
0%100%
200%
300%
C F F+P P
Annette Island � PAH Depletions
Annette % PAH depletion t1 to t4by treatment
-400%
-300%
-200%
-100%
0%100%
200%
300%
C F F+P P
Annette Island � PAH Depletions
C PF+PF
Barrow, t1-t4, %TPH Depletion
-80%-60%-40%-20%
0%20%40%60%
C F PF+P
Barrow - TPH DepletionsBarrow, t1-t4, %TPH Depletion
-80%-60%-40%-20%
0%20%40%60%
C F PF+P
Barrow - TPH Depletions
Barrow, t1- t4, %PAH Depletion
-20%
0%
20%
40%
60%
80%
C F F+P P
Barrow � PAH Depletions
Barrow, t1- t4, %PAH Depletion
-20%
0%
20%
40%
60%
80%
C F F+P P
Barrow � PAH Depletions
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Vegetated
Rhizosphere effect improves degradation�and
ti
tf
Control
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\HPC
HEM
\...\046R0101.D
Time (min.)
10 20 30 40
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\064R
0101.D
Time (min.)
10 20 30 40
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\067R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\070R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\077R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4Sig. 2 in E:\FID
54\080R0101.D
Time (min.)
T=1
Fall 1995
T=2
Spring 1996
T=3
Fall 1996
T=4Spring 1997
T=5Fall 1997
T=0
Summer 1995
T=5
Fall 1997
10 20 30 40
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\064R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\085R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\088R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\095R
0101.D
Time (min.)
10 20 30 400
1.0e4
2.0e4
3.0e4
4.0e4
Sig. 2 in E:\FID54\098R
0101.D
Time (min.)
10 20 30 400
10000
20000
30000
40000
Sig. 2 in C:\H
PCH
EM\...\052R
0101.D
Time (min.)
T=5
Fall 1997
T=1
Fall 1995
T=2
Spring 1996
T=3
Fall 1996
T=0
Summer 1995
T=4
Spring 1997
~700 days
Control diesel Spring 96
p.putida51%R.erthro
42%
unknown5%
M agilis2%
Control diesel Fall 96
p.putida52%
R.erthro18%
unknown14%
M.varians12%
B.cepacia4%
Control diesel Spring 97
p.putida45%
unknown21%
B.cepacia19%
R.erthro4%
A.ilicis4%
b.pumilis3%
S.prot3%
y.pseud1%
Veg/Fert diesel Spring 97
M varians18%
no match b15%
p.putida13%A.ilicis
9%
P.shigelloides6%
C.flaccumfaciens6%
d unknowns6%
match f5%
o match g5%
no match h5%
S.spiritivorum4%
R.aquatilis2%
M.mesophilicum2%
B.pumilus2%
R.fascians1%cyto.johnsonae
1%
Veg/Fert diesel Fall 96
p.putida53%
M.varians17%
Sphingobacterium (spiritivorum)
15%
Cytophaga (johnsonae)9%
unknow n4%
Curt. flacc2%
Veg/Fert diesel Spring 96
p.putida87%
S. malto8%
unknown5%
CulturableBacterialDiversity
Contaminant decreases are related to microbial shifts
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Culturable
Selective Media & FAME" CFUs/g" Identification of
culturable organisms
How can we characterize microorganisms in soil???
Phenotype
PLFA (Phospholipid fatty acids)
" Biomass" Community Composition" Physiological Status
Genotype
PCR (Polymerase Chain Reaction)t-RFLP (Terminal Restriction Fragment Length
Polymorphism) TGGE and DGGE (temperature/density gradient gel
electrophoresis)RT-PCR (real-time or quantitative PCR)
Enzyme
" Almost an expression of activity - Use DNA fragments for in situestimate of activity
" Soon � use RNA fragments for in situestimate of gene expression! enzyme synthesis
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Summary� It�s a system� We can adjust fertilizer and select plant species� Water and temperature more difficult to alter
Monitoring �??� �Standard� methods may not be appropriate� Multiple lines of evidence
� More �selective� chemical techniques� Possibly microbial techniques?
� The best response variable for monitoring may change with �status� of the system
� Phyto� Generally ≅ fertilizer initially� Phyto > fertilizer as recalcitrance ↑
� Fits well with concept of root-released carbon