bioremediation of land and water contaminated by toxic ... · chair professor of biology. city...

Bioremediation of Land and Water Contaminated by

Toxic Chemicals

Nora F.Y. TAMChair Professor of Biology

City University of Hong Kong29 October 2008

Eco Asia Conference, Hong Kong

Water and land contamination, a pressing world wide problem, poses a major threat to

Hong Kong and Mainland, particularly the rapid urbanization and industrialization cities

Common pollutants

●

Excess nutrients●

Heavy metals

●

Petroleum products●

POPs (persistent organic pollutants)

Halogenated compounds

DDT, PAHs, PCBs, PBDEs……

PAH emission

Oil pollution in marine environment Oil drill platform in Norway

1989 Exxon Valdez Incident, 42 Million litres of crude oil spreading in Alaska

524,000 litres of crude oil has leaked in the South China Sea some 15 km from the mouth of the Pearl River (Dec 2004)

Need to develop technologies

(i) Remove pollutants from wastewater and prevent them

enter the ocean

(ii) Clean-up contaminated environments

Treatment Technology●

Physical, chemical and biological (using bacteria) treatments are available

Ion exchange

Chemical precipitation

Filtration (membrane filtration)

Reverse osmosis

Activated sludge, biofilm

●

Crucial problemsCost?Generate secondary pollution (sludge)?Effective?

Alternate Technology

Do not generate secondary pollution problem

No sludge disposal

Ultimate solution

Simple, flexible and robust

Easy to set-up

Easy to operate

Cost-effective

Natural and environmental friendly

Provide Alternate Solution to Sewage and Contamination Problems

Human activities, e.g. waste

Contamination in land & water

Bio- and phyto- remediation of POPs and oil

Constructed mangrove for wastewater

Bisorption(Free and immobilized

microalgae)

Constructed Wetland Treatment

Popular for municipal wastewater and shrimp pond effluent in Europe, UK and North America

Increasing rapidly in Central and South America, Africa, Australia and New Zealand, and in Asia

Recent years, some constructed wetlands started in Asia

Comparison of conventional treatment and constructed wetland

Conventionaltreatment system

•

Involve large capital investment and operating costs

•

Require technical support

•

Small villages would not be able to afford such expensive conventional treatment systems

Constructed wetland system

•

Can be constructed where the wastewater is produced

•

Can be maintained easily, technical know- how is easy to grasp

•

Relatively lower-energy requirements and low- cost

•

Environmental friendly

Constructed Wetland Plants

●

Most CW use annual or perennial plants with frequent harvesting

Phragmites (common reeds)

Typha (cattails)

Scripus (bulrush)

Eichhornia (water hyacinths)●

Need frequent harvesting

Fluctuating removal efficiency

Labor intensive

Disposal of plant materials

Mangrove as constructed wetland for treating

wastewater

Sewage discharge directly

to river channel flooded

mangroves

Cattail vs mangrove plantPlant species Cattail MangroveTypes of wetland plant Emergent plant Emergent plantMaximum water depths found in wetland

treatment systems 0.1 to 0.75 < 0.05 to 0.5Flooding duration found in wetland treatment

(%) 70 to 100 50 to 100root to shoot ratio 0.4 to 0.6 0.7Anoxia endurance (days) >28 N/AAdaptation to high salinity No YesVegetative structures Rhizomes SeedlingRates of vegetation reproduction High MiddleNeeds of harvesting annually N/ACommunity dynamics invasive non-invasivePhysiological adaptations shallow water intertidalGrowth rate (kg dry wet ha−1 d−1) 21.92 to 167.12 80Annual nutrient uptake (nitrogen) (kg ha-1) 600-2630 159.92Annual nutrient uptake (phosphorous) (kg ha-1) 75-403 27.59

Mangrove wetlands

•

Similar to constructed wetland using cattails

•

No need to harvest annually•

Sediment is an efficient trap

•

Plants are robust and good in transporting oxygen to anaerobic sediments

•

Diverse microbes•

Adapt to alternate wet and dry condition, like intermittent sewage discharge

Electrical pump

Schematic Diagram of SSF constructed wetland

Effluent

influent

Gravel Sandy Soil

Plastic Septa

PVC column

Plastic box

Mangrove vs Cattails

Kandelia candel Typha latifolia

Compared with other CW studies (% removal municipal sewage)

Klomjek and Nitisoravut (2005)

Kaseva (2004)

Wild et al.

(2001)

Hong Kong(our study)

Cattail Sedge Cattail Cattail Cattail K. candel

HRT (d) 5 5 2.5 ND 5 5

BOD/OM 74.3 77.8 60.7 - 34.4 74.3

NH4 75.4 67.4 23.0 74.3 80.8 77.9

TKN - - - 40.3 65.6 65.5

P 44.9 35.6 - 74.8 27.2 97.2

Different mangrove plant species

Aegiceras corniculatum Avicennia marina Bruguiera gymnorrhiza (Bg)

Sonneratia caseolarisSonneratia apetalaKandelia obovata (Ko)

Acanthus ilicifolius (Ai)

Kandelia Aegiceras Mixed

DOC 78.07 78.60 75.31

PO4 98.00 98.40 97.40

NH4 80.12 88.19 86.68

Inorg N 56.80 57.15 65.08

TKN 55.49 57.04 55.00

Removal % by different mangrove species with 5-days HRT

Mangrove plants

Kandelia obovata

Aegiceras corniculatum

Sonneratia caseolaris

Species COD BOD5 TN NH3 -N TP SP

Influent from Futian (mg/L)

119.03 53.02 16.17 13.53 1.61 1.26

S. caseolaris 43.3564.9%

13.3875.5%

8.5653.6%

6.8752.6%

0.6565.0%

0.4569.2%

K. candel 41.9862.8%

13.7573.8%

8.2550.0%

7.2745.2%

0.6462.2%

0.4764.8%

A. corniculatum 37.7567.8%

13.6174.1%

7.9855.1%

6.0058.4%

0.4574.5

0.3276.9%

Concentration of effluent (mg/L）and removal % in 2 years (HRT 2.5 days)

% removal (HRT = 2.5 days)

Sonneratia Kandelia Aegiceras

Total bacteria

91.3 89.1 93.0

Coliform group

95.8 94.0 99.2

E. coli 95.5 90.0 99.8

Different types of wastewater●

Primarily settled municipal sewage

High organic matter, N and P removal

Plant uptake, microbial transformation, retention in sediment

●

Aquaculture pond effluent: e.g. shrimp

Inorganic N (300 mg L-1) >80% removal

Inorganic P (70 mg L-1): >99.8% removal

DOC (600 mg L-1): >98.5% removal

●

Livestock wastewater: e.g. pig farm

Inorganic N > 85%; P > 95% removal

●

Electroplating industrial effluent

Heavy metals: Cu, Zn, Cr, Ni and Cd

Accumulation in root, retention in sediment

Provide Alternate Solution to Sewage and Contamination Problems

Human activities, e.g. waste

Contamination in land & water

Bio- and phyto- remediation of POPs and oil

Constructed mangrove for wastewater

Bisorption(Free and immobilized

microalgae)

Bio- and phyto-remediation:

Can mangroves be used for remediation of POPs

(Persistent organic pollutants)?

Plant detectstoxicant

Chemical toxicant

Enhancedmineralizationof toxicant

Exudates stimulatemicrobial community

Change inroot exudation

Bioremediation: Plant-microbe-toxicant interaction

Inorganic Heavy metalsWetland plants: Spartina alterniflora; Phragmites australis

Weis and Weis, 2004

Organic Atrazine Tall fescue (Pennisetum clandestinum)

Singh et al., 2004

PAHsTree (Hibiscus tiliaceus) and vetiver (Vetiver zizanoides)

Paquin et al., 2002; Muratova et al., 2003

Crude oil Sorghum bicolorBanks et al., 2003

BTX Legumes (Galega orientalis)Suominen et al., 2000

PCBLegumes: alfalfa Grass: reed canarygrass, switchgrass and tall fescue

Chekol et al., 2004

Examples of Phytoremediation

Mangroves were planted in contaminated sedimentPAHsSpent lubricating oilKai Tat Approach ChannelVictoria Habour

Remediation of sediment contaminated with spent lubricating oil

0

2

4

6

8

10

3MAi IYAi Bg OC

TPH

-F3

mg/

g

0d

120d

% loss = Day 0 – Day 120 TPH•

Day 0 vs Day 120 TPH

41% 44% 36% 22%

Pyrene-contaminated sediment was placed at the surface (TS)

Pyrene-contaminated sediment was placed at the bottom (TB)

Flooded8 hrs day-1

Contaminatedsediment

Removal of pyrene (%)

Plant employed

K. candel B. gymnorrhiza

TS 96.42.5a 90.185.2b

TB 92.81.5a 85.86.6b

CN(no vegetation)

/

88.30.5b

PyrenePyrene removal: removal: KcKc > CN = > CN = BgBg ((pp = 0.05)= 0.05)More removal when contamination is in surfaceMore removal when contamination is in surface

Removal of pyrene from contaminated mangrove sediment

Remaining in sediment

Unlikely, very little amounts of pyrene added remained in sediments, also relatively immobile

Abiotic loss

Insignificant

Plant uptake

Mainly in roots but at low amounts

Microbial degradation in bulk sediment or in the rhizosphere

Major route to remove pyrene

Fate of Pyrene in mangrove microcosm

Bacterial colony grown on Phe-coated MSM agar (Arrow showing the clear zone)

KLHS SKOO

YOWG

HCCW

MWFG SPNY MPFG

PAH-degrading bacterial strains isolated from the enriched consortia

Strains *Source Bacterial name (% similarity)KLHS KLH Sphingopyxis alaskensis (97)SKOO SK Rhodococcus ruber (99)SKNG SK Sphingobium yanoikuyae (99)HCSS HC Rhodococcus opacus (98)HCCW HC Novosphingobium pentaromativorans (97)MWFG MW Sphingomonas sp. (98)YOWG YO Mycobacterium chlorophenolicum (99)SPNT SPN Paracoccus versutus (99)SPNY SPN Novosphingobium aromaticivorans (99)MPFG MP Sphingobium herbicidovorans (98)MPSS MP Sphingobium yanoikuyae (96)* The abbreviation of the source was the same as Fig.1

Bioaugmentation potential of different isolates in sediment slurry

H H 1 H H 2 H H 3 H H 4 H H 5 H H 6

0

2 0

4 0

6 0

8 0

1 0 0

Bio

degr

adat

ion

of P

AH

s (%

)

F l u o r e n e P h e n a n t h r e n e A n t h r a c e n e F lu o r a n t h e n e P y r e n e

H L 1 H L 2 H L 3 H L 4 H L 5 M S M S W

0

2 0

4 0

6 0

8 0

1 0 0

Bio

degr

adat

ion

of P

AH

s (%

)

Provide Alternate Solution to Sewage and Contamination Problems

Human activities, e.g. waste

Contamination in land & water

Bio- and phyto- remediation of POPs and oil

Constructed mangrove for wastewater

Bisorption(Free and immobilized

microalgae)

Biosorption

Retention of pollutants through passive adsorption, complexation and active absorption by biomass (biosorbents)Low costEfficient especially at low pollutant levelLittle or no secondary pollutionEnvironmental friendlyshare similar set-up as ion exchange

Micro- and macro-algae, fungi, and even non-living materials, e.g. rice husks, straws

Biosorption vs conventional treatment for heavy metals

Biosorption Precipitation Ion Exchange

Conc. dependence no yes no

Effluent quality (mg l-1) <1 2 - 5 <1pH adjustment rarely needed must sometime

sSelectivity good poor

(except Sulfide)

good

Efficiency good poor good

Regeneration good Not required good

Cost low high (Sludge) moderate

Microalgae as biosorbent

Ubiquitous and abundant

Rich biodiversity

Small size (in µm), large surface area to

volume ratio and high binding affinity

Fast growth and simple life cycle

Easy to culture and robust

Primary producers: autotrophic

Grow on carbon source: heterotrophic

Green and environmental friendly

Different microalgal species

Removal of heavy metals•

Screen most effective species

Isolates from sewage and polluted water

•

WW1 (Chlorella miniata) (HK)•

LC6A (Synechocystis sp.) (HK)

•

2f2aii, 2f2c, 3f2c, 4foa, 2f5aia (Chlorella sp.) (HK)•

3f2aiia, 3f2aiib (Cyanobacteria sp.)

•

C. sorokiniana 275 (Wuhan)•

Scenedesmus quadricauda 43 (Wuhan)

Commercial species•

Chlorella vulgaris

•

Selenastrum capricornutum

Removal of single heavy metal

Cu removal was usually highMost achieved >80% Chlorella species had >95% removal

Ni removal was generally lowerSome isolate (e.g. LC6A, 3f2aiia, 3f2aiib, 2f5aia)

had removal <20%Except WW1 (Chlorella miniata) achieved 80-

90%●

Zn, Pb and Cr removal varied among species

Repeated uses of two algae for Ni removal (10 cycles of 30 ppm Ni)

Treatment Cycle0 1 2 3 4 5 6 7 8 9 10

5

10

15

25

30

35

20

Ni c

once

ntra

tion

(µg/

ml)

C. vulgarisWW1

Reloading of fresh Nickel solution

Link

Metal adsorption capacity

Micro- algae

Cell diameter

(µm)

Specific surface

area (m2/mg)

Langmuir Qmax (mg/g cell)

Ni Cu Cr(III)

CV 3.39 0.018 12.06 18.72 ND

WW1 2.35 0.026 20.37 23.26 41.12

Qmax : maximum metal adsorption capacityCr(III) removal was due to bioreduction and biosorption

Removal of mixed heavy metals by microalgae

Algal species

Heavy Metals

Cr Zn Pb Ni Cu

Removal (%)

3f2aiib 48.6 ±

0.9 35.9 ±

15.7 35.2 ±

1.2 2.7±1.5 84.4 ±

1.1

2f5aia 70.2 ±

5.9 22.4 ±

4.0 86.5 ±

0.8 7.0±1.5 92.0 ±1.1

SC 72.9 ±

2.1 59.9 ±

6.0 79.3 ±10.7 27.0 ±

1.6 86.7 ±

1.3

3f2aiia 74.4 ±

1.7 22.5 ±

0.9 86.5 ±

0.5 5.3 ±

2.9 91.1±

0.4

Time required (min)

3f2aiib 1440 60 180 720 360

2f5aia 1440 90 1440 30 15

SC 120 360 60 120 5

3f2aiia 1440 720 1440 360 30

Removal by SC: Cu > Pb > Cr > Zn > Ni;Other pollutants like PAHs had little effects on metal removal

Mass culture in municipal wastewater

DW(24-0) > DW(16-8)

> BM(16-8) > DW(8-16)

Only little growth at

8-1ight-dark cycle

Municipal sewage is

good culture medium for microalgae

Production of Chlorella miniata biomass in settled municipal sewage

Kinetics of Cr(III) removal by different algal biomass (Cr(III) 100 mg L-1, initial pH 4.5)

Cr(III) biosorption

Cr(III) biosorption capacity by different algal biomass at various Cr(III) conc. (initial pH 4.5)

Free cells as biosorbent

Possible to obtain cheap microalgal

biomass from primarily settled municipal sewage with same removal efficiency

Algal production cost0.5 kg dw biomass per ton wastewater per day under outdoor condition with natural sunlightElectricity cost: HK$7.2 per day (based on 0.18 kw.h per ton wastewater)Production cost :HK$ 0.29 / kg dw biomass (compensate by the cost for treating primarily settled wastewater)

Immobilized algal beads●

Free cells are troublesome in operation, difficult to separate from treated effluent

●

Immobilization agents

Alginate

Carrageenan

Agarose

Chitosan

Polyacrylamide, etc.

Cd

day

0 2 4 6 8 10 12 14 16 18 20mg

met

al g

-1dr

y w

eigh

t mon

itor

0123456

chitosanagarosealginatepolyacrylamide

Metal adsorption and release profiles of biomonitors immobilized in different matrices (water Cd, Cu, Ni & Zn, all 1 mg L-1, for 10 days then metal free water for 10 days(mean and standard deviation; n = 3)

Ni

day

0 2 4 6 8 10 12 14 16 18 20mg

met

al g

-1dr

y w

eigh

t mon

itor

0123456Cu

day

0 2 4 6 8 10 12 14 16 18 20mg

met

al g

-1dr

y w

eigh

t mon

itor

0123456

Zn

day

0 2 4 6 8 10 12 14 16 18 20mg

met

al g

-1dr

y w

eigh

t mon

itor

0123456

Viability of algal beads after exposure to water containing mixture of Cd, Cu, Ni & Zn

Cells bead-1 Day 10

Alginate 2.9 x 105

Agarose 0

Carrageenan Bead dissolved after 2 days

Chitosan 0

Polyacrylamide 0

Immobilized microalgal beads

Carrageenan beads: dissolved easily

Polyacrylamide beads did not adsorb metals

Alginate beads adsorbed the highest amounts of metals, followed by chitosan and agarose

•

Calcium alginate: the most suitable immobilized agent

Production of alginate algal beads

Alginate microalgal suspension

Suspension pumped intoa Ca2+ solution

Microalgal and blank bead

2.5 x 106 cells bead-1

400 beads1 x 107 cells ml-1

Alginate immobilize microalgal beads (Bead diameter: 3-4 mm, Cell density; 106 cells bead-

Immobilization matrix

Algal cells(cell diameter: 3-6m)

Immobilized microalgal bead (diameter: 3-4mm)

T i m e ( m i n )

0 2 0 4 0 6 0 8 0 1 0 0 1 2 0 1 4 0 1 6 0

Res

idua

l Cu

leve

l in

solu

tion

(mg

L-1)

0

5

1 0

1 5

2 0

2 5

3 0

3 5

Cyclic Cu removal by algal beads (batch condition, shaking : 180 rpm, algal beads: 5.6 cells x 105

cells/bead, algal bead : metal soln

= 1:3 v/v)

reservoirof metalsolution

Algal beadscolumn

Peristalticpump

Effluent

Continuous Upflow

operation of the algal column

H y d r a u l i c l o a d i n g ( m l )

0 1 0 0 0 2 0 0 0 3 0 0 0 4 0 0 0

Efflu

ent c

oppe

r lev

el (m

g l-1

)

0

1

2

3

4

5

2 5

3 0

3 5

Influent Cu level

Effluent Cu level

Copper removal by algal columnContinuous upflow, HRT: 30 min

Time (hr)0 1 2 3 4 5 6 7

Cu desorbed (m

g L-1)

0

500

1000

1500

2000

2500

3000

3500

4000

Desorption of Cu after saturation study by 2% HNO3

Operation cycles

RT RT

T : Treatment Stage

R: Regeneration Stage

1st Cycle2nd Cycle

T i m e ( h o u r s )

0 2 0 4 0 6 0 8 0 1 0 0 1 2 0

Cop

per c

once

ntra

tion

(mg

L-1)

0

1 0

2 0

3 0

4 0

2 0 0

2 5 0

3 0 0

3 5 0

4 0 0

T R T R T R

Cyclic performance of algal column; T: Treatment R: RegenerationIn: Influent E: Effluent

In

E

Repeated uses of algal beads for removal of

TBT from wastewater

10 ppb

0 4 8 12 16 20 24

TBT

conc

. in

med

ium

(g

Sn L

-1)

0

5

10

15

20

50 ppb

0 4 8 12 16 20 24TB

T co

nc. i

n m

ediu

m (

g Sn

L-1

)

0

20

40

60

80

100 ppb

Time (days)

0 4 8 12 16 20 24

TBT

conc

. in

med

ium

(g

Sn L

-1)

0

30

60

90

120

150

180

1st 2nd 3rd 4th 5th 6th

Removal of PAH (polycyclic aromatic hydrocarbon): no difference between

free cells and algal beadsPyrene

0

20

40

60

80

100

0 50 100 150

Time (Hour)

% R

emov

al

Immobilized Suspended

Phenanthrene (3-ring PAH) Pyrene (4-ring PAH)

Phenanthrene

0

20

40

60

80

100

0 50 100 150

Time (Hour)

% R

emov

al

Immobilized Suspended

PAHs removal by SCFluorene

60

70

80

90

100

Day 1 Day 4 Day 7

% R

emo

val

Phenanthrene

60

70

80

90

100

Day 1 Day 4 Day 7

% R

emo

val

Fluoranthene

60

70

80

90

100

Day 1 Day 4 Day 7

% R

emov

al

Pyrene

60

70

80

90

100

Day 1 Day 4 Day 7%

Rem

ov

alBenzo[a]pyrene

60

70

80

90

100

Day 1 Day 4 Day 7

% R

emo

val

% removal of different PAHs by SC and effect of heavy metals

PAHs only PAHs + Metals

Microalgal beads bioreactor

Efficient biosorbent for heavy metals, POPs such as TBT and PAHs

Cheap as it can be produced from municipal sewage Algal production: <HK$0.3 per Kg dry biomassOperation: < HK$ 5 per ton wastewater

No secondary pollution problem, green technology

Higher flexibility Continuous, semi-continuous or batch columnsSame algal beads could be repeatedly used

Provide Alternate Solution to Sewage and Contamination Problems

Human activities, e.g. waste

Contamination in land & water

Bio- and phyto- remediation of POPs and oil

Constructed mangrove for wastewater

Bisorption(Free and immobilized

microalgae)

Thank you

E-mail: bhntam@cityu.edu.hk

Strain No. flo phe ant fla pyr Growth Isolated with 4-ring PAHsMycobacterium sp. HH1 2+++ 2+++ 7+ 2++ 2++ phe, pyrMycobacterium sp. HH2 3++ 1+++ - 1++ 1+ phe, pyrMycobacterium sp. HH3 3+ 1+++ - 3++ 8+ pheTerrabacter sp. HH4 3+ 2+ 7+ 1++ 14± flaTerrabacter sp. HH5 3+ 2+ 6++ 1++ 15± flaRhodococcus sp. HH6 2+ 2++ - 3+ - ngIsolated with 3-ring PAHsSphingomonas sp. HL1 1+ 1++ 4+ 1++ - pheSphingomonas sp. HL2 1++ 1++ 4++ 3± - pheSphingomonas sp. HL3 2+ 1++ 4+ - - pheSphingomonas sp. HL4 4++ 2++ 9++ 3+ - pheSphingomonas sp. HL5 2+ - - - - ng

Biodegradation of PAHs on R2A -PAH plates by the bacterial isolates obtained from HC sediment and their growth with PAHs as the sole carbon and energy source

Microalgal beads could degrade Pyrene but not blank alginate bead

Pyrene

0

20

40

60

80

100

0 50 100 150 200

Time (Hour)

% T

rans

form

ed /

Lost

Microalgal Bead Blank Bead

Pyrene

0

20

40

60

80

100

0 50 100 150

Time (Hour)

% R

emov

al

Microalgal Bead Blank Bead

Removal % Degradation %

Eutrophication and red tides due to excess nutrients

1998 Red tide in HK, killed >85% of culture fish; Loss > $300 M to mariculture industry;Unacceptable levels of toxins in shellfish

Persistent organic pollutants (POPs)CHCC lC lC l

C lC l

High level of pesticidesreported in human milk

ClCl

polychlorinated biphenyl

High level of PAHs and PCBs in marine mammals

Kinetics of Cr(VI) removal by different algal biomass (Cr(VI) 100 mg L-1, pH 2.0)

Cr(VI) removal by bioreduction and biosorption

Total Cr uptake by different algal biomass at various Cr(VI) conc. (initial pH 4.5)

Example of POPs: PAHs

PAHs (polycyclic aromatic hydrocarbons)

A group of aromatic compounds containing two or more fused benzene rings

Toxic, carcinogenic and mutagenic

16 PAHs: USEPA priority pollutants

Exist in various ecosystems, bind to soil and accumulate along food chains

Chlorella vulgaris

(commercial species)

WW1 C. miniata

(isolate from sewage)

Chlorella vulgaris

10 m

Chlorella miniata

10 m

Mangrove Wetland: complex and diverse biological system

Close to human activities, and has long been used as a convenient dumping site

Receive contaminants from tidal water, rivers, and land-based sources

Have an important role in pollution control through their absorptive capacity for organic pollutants and nutrients

Plants do not seem to suffer from waste discharge

T im e ( h r )0 1 2 3 4

Cu

deso

rbed

(mg

l-1)

0

5 0 0

1 0 0 0

1 5 0 0

2 0 0 0

2 5 0 0

3 0 0 0

0 2 4 60

2 0 0

4 0 0

6 0 0

8 0 0

Comparison of the desorption of Cu between the Algabiotor (20L) and the lab scale algal column (20 mL) (insert)

No scale up problem