Cadmium adsorption by mixed-culture biofilms
under metabolizing and non-metabolizing
conditionsJose Roberto Diaz
University of Puerto Rico at MayaguezAugust 3, 2006
Advisors: Dr. Robert Nerenberg Dr. Jeremy Fein
Mentors: Leon Downing Brian Ginn
Outline• Introduction• Objectives• Experimental method• Results and discussion• Conclusions• Future work• Acknowledgements
Introduction• Heavy metals are a
common cause of pollution
•non-degradable•accumulate in the
environment •causes of land and
water pollution • Conventional
treatment methods increasingly expensive heavy metals in the
atmosphere
Introduction• microorganisms treat heavy metal-
polluted wastes• accumulate trace levels of ions• major role in modification,
activation and detoxification• Immobilized systems have higher
surface areas and biological mass concentration
• ability of mixed cultures to adhere
• form biofilms• provide higher loading rates
than suspended systems
Introduction• Biofilms used in wastewater treatment systems
• resistant to inhibitory and toxic metals• tolerant to high metal concentrations• high affinity for metallic cations• anionic nature of polymers inhibit entrance of cationic
molecules to biofilm
• Immobilized-cell bioreactor technology provides cost-effective means for eliminating pollutants
Introduction• Live vs. Dead
Objectives
• Determine extent of Cd adsorption onto mixed-culture bacterial system under two conditions:
1. biofilm growth, metabolically active
2. biofilm growth, metabolically inactive
To waste
Experimental method
Acetate Feed = 16mM Phosphate buffer with 16.6356g CH3COOK (10 g/L acetate) to provide acetate as the electron donor
Continuous flow packed bed reactor
Q = 1mL/min
Retention time of 60 min
Air
N2
reactor is inoculated with Mishawaka activated sludge
Biofilm Growth Air is humidified to provide O2 as the electron acceptor
Experimental methodCd Adsorption Bulk solution:-no ammonium
-2 ppm Cd
-70 ppm Acet (dead)
-150 ppm Acet (live)
divide to test tubes
(5 mL each)
mix for approximately 2 hours
centrifuge and filter
add one-tenth of a gram of biofilm to each test tube
Cd in bulk liquid by
ICP-OES
Acetate by
Ion Chromatography
analyze samples
Cs-irradiation
35,000 rads/hr
Results and discussion
• Live or Dead?• Acetate consumption• Cd Adsorbed
Live or Dead?• metabolically active • metabolically inactive
Live-Dead staining
ImageLive
CountDead Count
Ratio (L:D)
1 1170 26 45.0000
2 944 19 49.6842
3 743 13 57.1538
4 1205 27 44.6296
5 710 11 64.5455
6 2579 41 62.9024
7 855 16 53.4375
8 577 13 44.3846
9 3214 91 35.3187
10 1835 57 32.1930
11 2727 57 47.8421
12 1598 28 57.0714
average 49.5136
stdev 10.0497
Image Live CountDead Count
Ratio (L:D)
1 346 96 3.6042
2 2175 1432 1.5189
3 311 186 1.6720
4 1038 87 11.9310
5 350 108 3.2407
6 71 85 0.8353
7 127 99 1.2828
8 187 107 1.7477
9 177 25 7.0800
10 93 61 1.5246
11 256 41 6.2439
12 530 621 0.8535
average 3.4612
stdev 3.3676
Metabolically Active BacteriaAcetate Control (150 ppm)
0
20
40
60
80
100
120
140
160
0 20 40 60 80 100 120 140
time (min)
Aceta
te Co
nc. (p
pm)
Run 1
Run 2
Acetate Consumption (live)20 ppm Acetate
05
1015202530
1 2 3 4 5 6 7
Sample#
Ace
tate
co
nsum
ed (p
pm)
20 ppm Acetate
18
20
22
24
26
28
1 2 3 4 5 6
Sample #
Ace
tate
co
nsum
ed (p
pm)
150 ppm Acetate
145147149151153155157
1 2 3 4 5 6 7 8 9 10
Sample #
Ace
tate
co
nsum
ed (p
pm)
Acetate Consumption (dead)50 ppm Acetate
0
20
40
60
80
1 2 3 4 5 6 7 8 9 10
Sample #
Ace
tate
co
nsum
ed (p
pm)
80 ppm Acetate
-100
-50
0
50
100
1 2 3 4 5 6 7 8 9 10
Sample #
Ace
tate
co
nsum
ed (p
pm)
70 ppm Acetate
0
20
40
60
80
100
1 2 3 4 5 6 7 8 9 10
Sample #
Ace
tate
co
nsu
med
(p
pm
)
Cd Adsorption for Metabolically Active Bacteria
0
5
10
15
20
25
30
35
40
45
1 2 3 4 5 6 7
Sample #
% C
d Ads
orbe
d 20 ppmAcetate20 ppmAcetate 2150 ppmAcetate
average %Cd adsorption = 26.9518
Cd Adsorption for Non-metabolically Active Bacteria
0
10
20
30
40
50
60
70
1 2 3 4 5 6 7 8 9Sample #
% C
d Ad
sorb
ed 50ppmAcetate80ppmAcetate70ppmAcetate
average % Cd adsorption = 48.9928
Cd Adsorption for Metabolically Active and Non-metabolically Active Bacteria
0
10
20
30
40
50
60
70
0 50 100 150 200
Acetate Consumed (ppm)
Cd A
dsor
bed (
%)
Active
Inactive
Discussion• bacteria are surviving the radiation
treatment• Non-metabolizing bacteria are still
consuming acetate in some samples more than others
• Metabolizing bacteria seem to be adsorbing less Cd (≈27%) than samples that have non-metabolizing bacteria (≈49%)
• possible reasons why
Conclusions• Radiation treatment is not effective in
killing bacteria and must be modified• Non-metabolizing bacteria certainly
showed an effect on how much Cd was adsorbed
• Samples with varying amounts of acetate adsorbed around the same percent of Cd for metabolizing and non-metabolizing bacteria
Future work
• Different method of killing bacteria• Non-metabolizing vs. metabolizing
in suspended growth system• Different electron acceptors and
donors for growing biofilm
Acknowledgements• EMSI• Dr. Valli
Sarveswaran• Leon Downing• Brian Ginn• Dr. Jeremy Fein• Dr. Robert
Nerenberg• CEST