Phytoremediation of Arsenic Phytoremediation of Arsenic Contaminated Soils Using Contaminated Soils Using

Chinese Brake FernChinese Brake Fern

Maria Silva

Soil and Water Science Department

University of Florida

ArsenicArsenic Anthropogenic activities account for most As

contamination in soils and water;

As is carcinogenic

Remediation of As contaminated sites has become an important issue

Current remediation technologies are expensive and environmental disruptive.

The use of plants to remove, contain, or render harmless environmental pollutants.

PhytoremediationPhytoremediation

Plant biomass

Uptake

Plant species

Contaminant concentration

Use of accumulating plants to remove metals from soil by concentrating them in

harvestable parts.

PhytoextractionPhytoextraction

Chinese Brake fern (Pteris vittata L)

Natural ability to tolerate, accumulate, and translocate high concentrations of As

As high as 2.3% in fronds

Rapid growth rate

Perennial

High biomass

Capable of taking up both organic and inorganic As

Although the fern shows great

potential to be used in

Phytoremediation, many

questions still need to be

answered!!!

Research objectivesResearch objectives1. Determine the effectiveness of Chinese

Brake fern (CBF) in continuously remove As from soils over time;

2. Investigate the impacts of CBF on As mobilization and redistribution in the rhizosphere soil

3. Determine the NPK levels for optimum As removal by the fern

4. Evaluate the effect of plant maturity on As accumulation by the fern.

Experiment 1Experiment 1Effectiveness of Chinese Brake fern on

arsenic removal over time

HYPOTHESESHYPOTHESES:

1. Chinese Brake fern is capable of continuously removing As from soils;

2. Arsenic availability in soil will decrease as more available As is taken up by Chinese Brake

3. Changes in plant available As can be predicted by partitioning the soil As-pools using a sequential fractionation procedure

Materials and MethodsExperimental setup

Completely Randomized Design Six As-contaminated soils One plant per pot containing 4 kg soil Four replications Controls without plants

Sampling Harvest fronds 2-3 times a year Collect soil with plant harvest

Soil

Source of arsenic As (mg kg-1)

Marl Natural 20

Avon Cattle-dipping vat 27

CCA chromated-copper-arsenate

(CCA) wood preservative

110

CDV Cattle-dipping vat 300

Mining Mining activities 300

EDS herbicide 700

Experimental Soils

Analyses

Aboveground biomass

Total As concentrations (soil and plant)

Hot block digestion system (EPA Method 3050A)

Graphite furnace AAS

Arsenic fractionation in soil

Wenzel et al. (2001)

Fractions Extracting solution Extraction condition

SSR*

As-NAs-N: Non-specifically- bound

(NH4)2SO4 0.05M 4h shaking, 20oC 1:25

As-SAs-S:Specifically-bound

(NH4)H2PO4 0.05M 16h shaking, 20oC 1:25

As-AAs-A:Amorphous hydrous oxide-bound

NH4-oxalate buffer

(0.2 M) pH 3.25

4h shaking, 20o dark

1:25

As-CAs-C:Crystalline hydrous-oxide-bound

NH4-oxalate buffer

(0.2 M) + ascorbic acid (0.1M) pH 3.25

30 min in a water basin at 96oC with light

1:25

As-RAs-R:Residual HNO3/H2O2 Hot block digestion

1:50

SSR=soil to solution ratio

As fractionation procedure

As availab

ility

Experiment 2 Arsenic mobilization and redistribution

in the rhizosphere of two ferns

HYPOTHESESHYPOTHESES:

1. More roots of CBF will be developed in arsenic-rich soil than arsenic-free soil;

2. Changes in the rhizosphere of CBF will be greater than those of Boston fern, a non-hyperaccumulator.

Materials and MethodsExperimental setup

Completely Randomized Design

Factorial scheme (2 x 2)

2 plants (CBF and Boston fern)

2 soils (contaminated and non-contaminated)

1 plant/pot containing 2.5kg of soil

4 replications

Controls without plantsSampling

Harvest after 8 weeks of growth

Collect rhizosphere and bulk soil

Rhizopot

Rhizosphere soil

Bulk soil

45um

AnalysesPlant

Frond and root biomass Frond and root As concentration Root length density Root area density

Soil (rhizosphere and bulk) water-soluble As Total As pH and DOC As fractionation

Experiment 3Interactive effects of N, P, K and As on

plant arsenic uptake

HYPOTHESESHYPOTHESES:1. As accumulation affects plant nutrient

requirements;

2. Maximum plant As removal can be achieved through optimum application of NPK;

Materials and MethodsExperimental setup

4 factor- 5 level central composite design

N = 60 to 300 ppm;

P = 30 to 150 ppm;

K = 30 to 300 ppm;

As = 50 to 200 ppm;

one plant/pot with 1.5 kg of soil

Three replications

Sampling Harvest after 8 weeks of growth

Soil

Analyses

Plant Frond and root biomass Frond and root As concentration Frond and root macro and micronutrients

Soil (rhizosphere and bulk) Total As concentrations pH and DOC

Experiment 4Arsenic removal by Chinese brake fern

using plants of different maturity

HYPOTHESESHYPOTHESES:

1. Plant maturity affects its arsenic removal capability;

2. Maximum plant As removal can be achieved using optimum plant age;

Materials and Methods

Experimental setup Completely Randomized Design

One plant per pot with 2.5 kg of soil

Plants: 45 days, 8, 10 and 16 month-old

One As contaminated soil (150 mg kg-1)

Four replications

Sampling Harvest after 12 weeks of growth

Bulk and rhizosphere soil

Plant biomass

As concentrations in soil and plant

Soil pH and DOC in the rhizosphere and

bulk soil

Analyses

Preliminary Results

0

100

200

300

400

500

Marl Avon CCA Mining CDV EDS

Soils

Biom

ass

incr

ease

(%

)

Frond biomass in the first harvest

Initial frond biomass = 8g

Experiment 1

Fern As concentration in the first harvest

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

Marl Avon CCA Mining CDV EDS

Soils

As(

mg

kg-1

)

Arsenic removed from soils by fern after one harvest

0

1

2

3

4

5

6

7

8

9

10

Marl Avon CCA mining CDV EDS

Soil

%o

fAs

rem

ov

ed

Changes in Soil Arsenic fractions after first harvest

As-N non-specifically bound, As-S specifically bound, As-A amorphous Fe and Al As-C crystalline Fe and Al As-R residual

0%

20%

40%

60%

80%

100%

Soils

As

As-R

As-C

As-A

As-S

As-N

Clean soil

As soil

Experiment 2

Clean soil

As soil

Chinese brake fern

Boston fern

Plant biomass, As concentration & removal after 8 weeks of growth

 Fern specie

Biomass As concentration As remediated

(%)Frond Root Frond Root

--- g plant-1--- -- --mg kg-1 -----

  Non-contaminated soil

CBF 6.33 1.98 0.58 2.65 -

Boston 2.46 0.86 0.10 0.30 -

Contaminated soil

CBF 5.54 1.84 432 116 1.03

Boston 1.75 0.60 19.7 98.0 0.04

0

0.5

1

1.5

2

C. Brake Boston No plant

Treatment

W-S

As

(mg

kg-1

)

Bulk soil Rhizosphere

Water-soluble As

a a a

C BA

6.9

77.1

7.27.3

7.47.5

7.67.7

C. Brake Boston No plant

Treatment

So

il p

H

Bulk soil Rhizosphere

Soil pH

a a a

A

B B

Summary-1

1. CBF grew well in all six soils with 195-360% biomass increase after 4 months;

2. CBF was effective in removing As from the soils, with 4- 9% reduction after one harvest;

3. The majority of soil As was associated with the amorphous fraction

Summary-2

4. Compared to Boston fern, CBF accumulated greater biomass and greater arsenic in the fronds;

5. Water-soluble As was lower in the rhizosphere of CBF than Boston fern;

6. pH was greater in the rhizosphere of CBF than Boston fern;

Special Thanks

Advisor- Dr. Lena Ma

Dr. Jorge Santos

Committee members:

Dr. Commerford

Dr. Rhue

Dr. Stamps

Dr. Bonzongo

Tom Luongo

Trace metal biogeochemistry group

Sponsorship – CAPES

Thank you