growth inhibition by soil components for degradation of dioxins using white rot fungus

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

Growth inhibition by soil components

for degradation of dioxins using white rot fungus

11

Shinya SuzukiShinya Suzuki, , Ayako Tachifuji, Yasushi MatsufujiAyako Tachifuji, Yasushi Matsufuji

Department of Civil Engineering, Fukuoka University8-19-1 Nanakuma, Johnan-ku, Fukuoka, 814-0180 JapanE-mail: ssuzuki@fukuoka-u.ac.jpssuzuki@fukuoka-u.ac.jp

O

Cl

Cl

O

OCl

O

Cl

Cl

Cl

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

1-1 Background in Japan1) Dioxin compound

- risk to cause long-term contamination in soil- chemically stable structure- accumulate in environment

2) There are still many contaminated soils- not only around MSW incineration facilities - but also around rice field

- included in pesticide as impurities about 40 years ago

3) Physicochemical treatment methods- already developed, but- disadvantage from economic and energetic point of view

1) Dioxin compound- risk to cause long-term contamination in soil- chemically stable structure- accumulate in environment

2) There are still many contaminated soils- not only around MSW incineration facilities - but also around rice field

- included in pesticide as impurities about 40 years ago

3) Physicochemical treatment methods- already developed, but- disadvantage from economic and energetic point of view

4) So, for remediation of contaminated soil - which has low concentration of dioxins- more effective to use bioremediation method

4) So, for remediation of contaminated soil - which has low concentration of dioxins- more effective to use bioremediation method

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

1-2 Objective

Objective of this study- to accomplish effective bioremediation method

- for low levels of dioxins contaminated soil- by using “Phlebia brevispora”

- to clarify inhibitory factor of soil components - to evaluate relationship between growth of fungi and degradation of dioxins

Objective of this study- to accomplish effective bioremediation method

- for low levels of dioxins contaminated soil- by using “Phlebia brevispora”

- to clarify inhibitory factor of soil components - to evaluate relationship between growth of fungi and degradation of dioxins

In particular, - method using white rot fungi attracts attention- but living spheres of such fungi are usually in dead trees, fallen trees, etc.

It is still unclear whether - such fungi can survive and degrade dioxins in soil environment.

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

Soils: composed from various materials such as clay minerals, organic substances

- to compare difference of degradability according to organic substances

Soils: composed from various materials such as clay minerals, organic substances

- to compare difference of degradability according to organic substances

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

2-1 Materials

Organic-poor soil Organic-rich soil

2,7-DCDD- degraded easily, shorter incubation period- simplified extraction

O

Cl

Cl

O

1,3,6,8-TCDD-main components of CNP/PCP originating dioxins- longer incubation period

OCl

O

Cl

Cl

Cl

Phlebia brevispora (TMIC 33929)- can degrade 1,3,6,8-TCDD- maintained on potato dextrose agar (PDA) medium

Phlebia brevispora (TMIC 33929)- can degrade 1,3,6,8-TCDD- maintained on potato dextrose agar (PDA) medium

unitorganic-poor

soilorganic-rich

soil

ｐＨ(CaCl2) - 4.3 5.4

Ignition Loss (%) 4.0 12.8

TOC mg/g 0.4 31.7

TN mg/g 0.1 3.1

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

2-2 Experimental condition2,7-DCDD 1,3,6,8-TCDD

no soil ○ ○- -

organic-poor soil ○ -(L/S=6) -

organic-rich soil ○ ○(L/S=6) (L/S=6)

○(L/S=9)○

(L/S=12)○

(L/S=15)

Influence of soil property- L/S=6

Influence of Liquid-Solid (L/S) ratio- 1,3,6,8-TCDD- organic-rich soil

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

2-3 Analytical procedurePre Incubation

-Kirks HCLN medium: 30mL(autoclaved)

-Fungus body- Incubation: 14 or 30days-Temp.: 25℃-shaking every time

Growth of fungi“Weight analysis”

Growth of fungi“Weight analysis”

Degradation of dioxins- “2,7-DCDD”- “1,3,6,8-TCDD”

Degradation of dioxins- “2,7-DCDD”- “1,3,6,8-TCDD”

-Incubation: 5days-Temp.: 25℃

Incubation

Analysis (n=3)Mixing

or-Distilled water: 15mL(L/S = 9)

30mL(L/S = 12)45mL(L/S = 15)

no addition0mL(L/S = 6)

Simulation of contaminated soil

Organic-poor soil: 5g (autoclaved)

-2,7-DCDD

Organic-rich soil: 5g (autoclaved)

or

-1,3,6,8-TCDD

or

2,7-DCDD or 1,3,6,8-TCDD

Liquid-state condition

Slurry-state condition

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

0

20

40

60

80

100

120

no soil2,7-DCDD

organic-poorsoil

2,7-DCDD

organic-richsoil

2,7-DCDD

reco

very

rat

e of

2,7

-DC

DD

(%)

control

treatment

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-1-1) Influence of soil property L/S = 6

O

O Cl

Cl O

O ClCl

ClCl

O

OCl

Cl

OH

O

OClCl

ClCl

OHOH

O

OCl

Cl

OCH3

O

OClCl

ClCl

OCH3OCH3

2,7-DCDD

- monohydroxy-DCDD, monomethoxy-DCDD were obtained as metabolite

- monohydroxy-DCDD, monomethoxy-DCDD were obtained as metabolite

Degradation rate of 1,3,6,8-TCDD: only 1.1% in slurry-state with organic-rich soil

Degradation rate of 1,3,6,8-TCDD: only 1.1% in slurry-state with organic-rich soil

76.3% 66.1% 12.7%

0

20

40

60

80

100

120

no soil1,3,6,8-TCDD

organic-richsoil

1,3,6,8-TCDD

reco

very

rat

e of

1,3

,6,8

-TC

DD

(%)

control

treatment

35.0%1.1%

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-1-2) Liquid-solid ratio 1,3,6,8-TCDD in organic-rich soil

Dissolved matters from organic-rich soil had inhibition activity?- this improvement probably caused by dilution of inhibition material in the soil ?

Dissolved matters from organic-rich soil had inhibition activity?- this improvement probably caused by dilution of inhibition material in the soil ?

0

20

40

60

80

100

120

L/S=6 L/S=9 L/S=12 L/S=15

reco

very

rat

e of

1,3

,6,8

-TC

DD

(%)

control

treatment

1.1% 18.9% 32.2% 27.0%

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-2-1) Soil extractionSo, in order to examine inhibition activity of soluble matters only

- organic-rich soil was washed in advance to fractionate into…,So, in order to examine inhibition activity of soluble matters only

- organic-rich soil was washed in advance to fractionate into…,

organic-rich soilorganic-rich soil dissolved matterdissolved matter

washed organic-rich soilwashed organic-rich soil

itemsoil growth amount degradation rate

no soil ○ ○- -

organic-rich soil ○ ○(L/S=6) (L/S=6)

2,7-DCDD

dissolved matter ○ ○- -

washed organic-rich soil ○(L/S=6)

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-2-2) Analytical procedurePre Incubation

-Kirks HCLN medium: 30mL(autoclaved)

-Fungus body-Incubation: 14days-Temp.: 25℃-shaking every time

Growth of fungi“Weight analysis”

Growth of fungi“Weight analysis”

Degradation of dioxins- “2,7-DCDD”

Degradation of dioxins- “2,7-DCDD”

-Incubation: 5days-Temp.: 25℃

Incubation

Analysis (n=3)

2,7-DCDD Extraction

-24hours-shaking

Solvent: distilled water

Only soluble matters from organic-rich soil were added into flask

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

0.00

0.05

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0.30

0.35

0.40

0.45

0.50

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100

pre incubationDay 0

incubationDay 0

incubationDay 10

incubationDay 14

grow

th a

mou

nt o

f fun

gi (

g)

degr

adat

ion

rate

of 2

,7-D

CD

D (%

)

incubation time

no soil

dissolved matter

organic-rich soil

no soil

dissolved matter

organic-rich soil

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-2-3) Growth and degradation rate

In case of dissolved matters, growth amount of fungus: much smaller- Growth inhibition was clearly confirmed during pre-incubation period

In case of dissolved matters, growth amount of fungus: much smaller- Growth inhibition was clearly confirmed during pre-incubation period

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-2-4) Relationship growth amount v.s. degradation rate

Degradation rate of dioxins in liquid-state only depended on growth amount- soluble matters from organic-rich soil had growth inhibition activity,- it was a part of whole inhibitory effect from organic-rich soil.

Degradation rate of dioxins in liquid-state only depended on growth amount- soluble matters from organic-rich soil had growth inhibition activity,- it was a part of whole inhibitory effect from organic-rich soil.

y = 293.01xR² = 0.917

0

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50

60

70

80

90

100

0.00 0.05 0.10 0.15 0.20 0.25 0.30

degr

adat

ion

rate

of 2

,7-D

CD

D (%

)

growth amount of fungi (g)

no soil

dissoleved matter

organic-rich soil

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

0

20

40

60

80

100

120

organic-rich soil2,7-DCDD

washed organic-richsoil

2,7-DCDD

no soil2,7-DCDD

reco

very

rat

e of

2,7

-DC

DD

(%)

control

treatment

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

3-2-5) Influence of growth inhibition

Difference of degradation rate of 2,7-DCDD- between “organic-rich soil” and “no soil” was about 60%, - growth inhibition: about 35%, other inhibition: about 25%

Difference of degradation rate of 2,7-DCDD- between “organic-rich soil” and “no soil” was about 60%, - growth inhibition: about 35%, other inhibition: about 25%

25%

35%

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

4-1 Conclusions Growth inhibition by soil components

For degradation of dioxins using white rot fungusGrowth inhibition by soil components

For degradation of dioxins using white rot fungus

2) Organic-rich soil has large inhibition activity- soluble matters has growth inhibition effect

2) Organic-rich soil has large inhibition activity- soluble matters has growth inhibition effect

1) Degradation rate of 1,3,6,8-TCDD: - only 1.1% in case of organic-rich soil- increased according to change liquid-solid ratio

1) Degradation rate of 1,3,6,8-TCDD: - only 1.1% in case of organic-rich soil- increased according to change liquid-solid ratio

3) Difference of degradation rate of 2,7-DCDD between with and without “organic-rich soil”: about 60%- of this, growth inhibition accounted for about 35%

3) Difference of degradation rate of 2,7-DCDD between with and without “organic-rich soil”: about 60%- of this, growth inhibition accounted for about 35%

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

1. Introduction 2. Materials and Methods 3. Results and Discussion 4. Conclusions

4-2 Conclusions

In order to establish remediation method for soil- to important to consider soil components- to control water content

In order to establish remediation method for soil- to important to consider soil components- to control water content

- Water content: most important factor in remediation of contaminated soil- This improvement in degradation of dioxins

- probably caused by dilution of growth inhibition material in soil- In fact, degradation of dioxins increased by soil washing in advance

- Water content: most important factor in remediation of contaminated soil- This improvement in degradation of dioxins

- probably caused by dilution of growth inhibition material in soil- In fact, degradation of dioxins increased by soil washing in advance

Shinya Suzuki, Fukuoka University, Japan. Sardinia 2011

Thank you very much for your attention !Thank you very much for your attention !