Biological Den. 1

8/8/2019 Biological Den. 1

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Biological DenitrificationIn Aquaria

Prepared by:068044 Merve Ayvaz

October 27, 2009


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OUTLINE

Part 1 Overview to the Biological

Denitrification

Part 2 Principles of Denitrification

Methods

Part 3 Comparison of ACD and HCDMethods on an Experiment


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PART 1

OVERVIEW

TOBIOLOGICAL

DENITRIFICATION


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Nitrate Accumulation

Side effects;

o Direct toxicity to organisms

o Alkanity and pH effect

o Decrease in oxygen concentration


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Nitrate Accumulation

Regular water exchange is impractical for large aquaria.

Biological denitrification has been selected as a method of

choice for controlling NO3

- concentration in closed systems.


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What is Biological Denitrification?

Reduction of nitrate to nitrogen gas in suboxic

conditions by bacteria.


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DenitrifyingBacteria

Denitrifying bacteria is a part of nitrogen cycel

Facultative aerobes

Enzymatic reduction


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DenitrifyingBacteria

Thiobacillus denitrificans

Micrococcus denitrificans/Paraoccus denitrificans

Pseudomonas


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PART 2

PRINCIPLES OF

DENIT

RIFICAT

IO

NMETHODS

ACD &HCD


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Methods for Biological Denitrification

Autotrophic columnar denitrification (ACD)

Heterotrophic columnar denitrification (HCD)


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Heterotrophic and Autotrophic Columnar

Denitrification

Heterotrophic

means obtaining

ready made organic

food from the

environment

Autotrophic means

manufacturing food

from inorganic

compounds usually

carbon dioxide and

water.


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Heterotrophic and Autotrophic Columnar

Denitrification

HCD: Heterotrophic bacteria + Organic carbon source as a

food and electron donor

ACD: Autotrophic bacteria + Inorganic carbon source tomanufacture food and sulfur as electron donor

In both cases it is essential to provide an anaerobic

environment within the biofilter in order to allow thereduction process to start.


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HCD, Heterotrophic Columnar

Denitrification

Oxidation of different organic matters using while the

reduction of NO3- to N2 as the electron acceptor.

Carbon source has to be added periodically to promote

growth and metabolism of heterotrophic bacteria.

2

-

32

275246126

-

3

0.86CO0.38HCOO1.62H

ONH0.62C0.5N0.62NHOH0.72CNO1)

p

-

2223

-

3 6OHO7H5CO3NOH5CH6NO2) p


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Pseudomonas Bacteria

Gram-negative rod bacteria

Commonly found in soil, ground water, plants and animals

T

here are two type ofP

seudomonas bacteria,P

seudomonasstutzeri and Pseudomonas aeruginosa

Pseudomonas stutzeri produced only dinitrogen;

Pseudomonas aeruginosa produced nitrous oxide as well


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ACD, Autotropic Columnar Denitrification

The chemoautotrophic bacteria Thiobacillus denitrificans

oxidize sulphur and sulphur compounds while reducing NO3-

to free dinitrogen gas (N2).

In this case, the metabolic reaction would progress according

to;

p

9.62H5.4N(biomass)NOH0.92C

11SOO2.5H1.71NH0.5CO4.1HCONO1011S

2275

-2

4242

-

3

-

3


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Thiobacillus Denitrificans

Gram-negative, colorless, rod-shaped bacteria

Thiobacillus are obligate autotrophic organisms, meaning they

require inorganic molecules as an electron donor and inorganic

carbon (such as carbon dioxide) as a source.

They obtain nutrients by oxidizing iron and sulfur

with O2 or NO3-.


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PART 3

COMPARISON

OFACD & HCD METHODS

ON AN EXPERIMENT


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Aim of the Experiment

The nitrate removal rates for ACD and HCD methods will

be compared

pH effect and reversibility of the reaction will be discussed

Control Parameters

Temperature is constant at 23 and 26 oC

Salinity, pH, dissolved oxygen, temperature are followed in

each set


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Experimental Set-up

PUMP

Glucose

injection

Denitrification

column

Outflow

Flow

regulatio

n


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Experimental Set-up

The pilot plants had a total workingvolume of 50 L with a

circulating flow system between the glass holding tanks and

the denitrification unit.

It is a PVC cylinder with an internal diameter of 8 cm,

packed with specific media for conditioned bacteria fixation

to a height of 40 cm.


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Experimental Set-up

Upward circulation of water was chosen to ease the

evacuation of gaseous nitrogen via the open top end of the

column.

o Fixation of heterotrophic bacteria was induced by periodicaladdition of an organic carbon source (1 ml of 20% glucose

aqueous solution).


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Packing Materials for Column

The most studied packing materials are stones, clay, schist,

and plastic of various types, such as polyethylene,

polyesterene and even waste plastic materials.


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Packing Materials in Column

An adequate packing media should be;

inexpensive

easily available

high area/volume ratio

good mechanical resistance

suitable for microorganism attachment

Among natural packing materials, volcanic stones reasonablymeet these criteria due to its high natural porosity, relative

resistance and inert behavior.


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Packing Materials in HCD Column

HCD: Porous volcanic media with 7 mm particle size.Volcanic rocks mostly used medium in aquarium, pond

and marine denitrification systems.o Pseudomonas bacteria attaches inherently to the volcanic

rocks porous surface.


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Packing Materials inACD Column

ACD: Elemental sulphur (commercial powder) with 24 mm

particle size was used as bacterial fixating media.

The mechanism by whichThiobacillusDenitrificans can use solid-phase

electron donors thatcannot betaken

into thecell is ofconsiderable interest

butthat is currently unknown.


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Start of the Experiment

An artificial seawater solution (salinity = 36%) was prepared

with synthetic salt diluted into freshwater.

Artificial seawater that was used in the pilot study,purposefully contaminated with nitrate by the help of

nitrifying bacteria and NH4Cl.

When adequate nitrate concentration was present (after the

nitrification process) bicarbonates were added to raise pH to

8.4 and create a suitable environment for denitrification to

progress.


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Crucial Points for Denitrification

Nitrate concentration

Organic carbon source supply

Oxygen concentration, pH,


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Water Supply to the Column

Water flow was kept low (1.5 dm3/h) in the beginning to

accelerate the creation of an anaerobic environment in the

column for both methods.

Oxygen resistance of tolerance of autotrophic bacteria is

higher then heterotrophic bacteria. Water flow rate raised

after nitrated reduction started for ACD.


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RESULTS

HCD ACD


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RESULTS

HCD ACD

p

9.625.4(biomass)0.92

112.51.710.54.11011

2275

-2

4242

-

3

-

3


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Results,HCD


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pH Effect in HCD


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Results,ACD


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pH Effect in ACD

Bicarbonate addition in day 60

Suggested pH for T. Denitrificants

is 6.2


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Conclusion

ACD showed double nitrate reduction rates then HCD

Results are under effect of microbiological group that

produce N2 depending on adequate pH and oxygen resistant

Inorganic carbon has great control on ACD

Organic carbon has chief control on HCD


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THANK YOU FOR

YOUR ATTENTION

Biological Den. 1

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