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BIODEGRADATION- OVERVIEW & BIODEGRADATION OF HYDROCARBONS & PCB Gunjan Mehta Deptt. of Biotechnology, Virani Science College, Rajkot

Biodegradation overview

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Page 1: Biodegradation overview

BIODEGRADATION- OVERVIEW &

BIODEGRADATION OF HYDROCARBONS & PCB

Gunjan Mehta

Deptt. of Biotechnology,

Virani Science College, Rajkot

Page 2: Biodegradation overview

Degradation

Degradation is breakdown of complex organic material into simpler one.

Different ways of degradation: 1. Photodegradation by natural day light 2. Oxidation by chemical additives(Catalysts) 3. Thermal degradation by heat 4. Mechanical degradation by mechanical

force 5. Biodegradation by Microorganisms.

Page 3: Biodegradation overview

Degradation

Three levels of degradations: 1. Rapid degradation(day- week): HC

compounds

2. Slow breakdown(Months- years): HC polymers/ Halogenated compounds

3. No degradation: Recalcitrant/ Xenobiotic: Plastic

Page 4: Biodegradation overview

Biodegradation

“Natural and complex process of decomposition facilitated by biochemical reactions.”

It is biological transformation of an complex organic material to simpler by Mos.

Reduced organic materials are thermodynamically unstable and oftenly attacked by microbial enzymes.

Biodegradibility: Quality, representing the susceptibility of the substrate to biological transformation.

Natural recycling of matter which mediated by large consortium of Mos.

Page 5: Biodegradation overview

Types of Biodegradation

1. Primary biodegradation: Biochemical ways of catalysts where

transformation or alteration in chemical structure of a compound occurs by biochemical reactions.

Results in loss of specific property- partial biodegradation and leaves molecule mostly intact.

Not desirable due to toxicity issues. Ex. Change in toxic halogen gp from Pera to

Meta position. Azo dye Amino benzene

Page 6: Biodegradation overview

Types of Biodegradation

2. Acceptable biodegradation: Biological conversion of toxic compounds to

non toxic by biological means. Removal of undesirable characteristics

occurs. Complete removal of toxic entity occurs.

Page 7: Biodegradation overview

Types of Biodegradation

3. Ultimate biodegradation: The level of degradation where the

compound is totally utilized and results in production of CO2 and water and mineral constituents.

Molecular cleavage is so extensive that it removes all chemical, biological and toxic properties.

The ultimate products are highly stable and can’t be degraded further.

Ex.

Page 8: Biodegradation overview

Reactions involved in Biodegradation Oxidative reaction Reductive reaction Hydrolytic reaction (water) Conjugative reaction (Methylation,

Acetylation)

Page 9: Biodegradation overview

Factors affecting Biodegradation

Substrate

related

Organism

related

Environment

related

Page 10: Biodegradation overview

Factors affecting Biodegradation

Sub

stra

te

rela

tedNature of pollutants

Physiochemical properties

Concentration

Biodegradability

Toxicity

Chemical nature

Volatility

Polarity

Page 11: Biodegradation overview

Factors affecting Biodegradation

Org

an

ism

re

late

dPopulation density

CompositionIntra/ Inter specific

interactionEnzyme activity

Turn over number

Adaptation

Page 12: Biodegradation overview

Factors affecting Biodegradation

En

vir

on

men

t re

late

d

Temperature

pH

Oxygen availability

Nutrient sources- C & e-

Salinity

Page 13: Biodegradation overview

Organisms responsible for biodegradation

Most significant group of living organism involved in biodegradation, responsible for 65% total metabolism due higher growth rate and biomass. Higher organism are also involved but not significantly, inability to degrade complex molecule.Microbes represent most diversified metabolism on earth.

Microbes--------Complex material-----simpler form

Microbes utilize energy more efficiently in comparison to higher organisms.

High rate of reproduction and mutation is the governing factor.

Page 14: Biodegradation overview

Organisms responsible for biodegradation

Other lower organism- algae and invertebrates too possess some of the criteria- Earthworm, but their biodegradative potential is still unknown.

Marine biodegraders: Bacteria and Fungi Soil biodegraders: Bacteria and Fungi Mutations are very often in bacteria and is

very useful for progressive adaptation towards the biodegradation pathways.

Not all microbe are equipped with all enzyme, so many of them follow….COMETABOLISM

Page 15: Biodegradation overview

Organisms responsible for biodegradationIm

port

ant

bact

eri

aAchromobacter, Acinetobacter

Alcaligenes, Arthrobacter

Bacillus

Flavobacterium, Nocardia

Pseudomonas spp.

Page 16: Biodegradation overview

Reactions involved in Biodegradation

Three categories of biodegradation:I. Usable immediately II. Usable following acclimatization III. Recalcitrant IV. Cometabolic reaction

Page 17: Biodegradation overview

Reactions involved in Biodegradation I. Usable immediately Simple sugars, amino acids and fatty acids-

direct utilization. The enzymes required for breakdown are

either constitutive or inducible. This requires minimum acclimatization period.

II. Usable following the acclimatization: A lag phase is required for adaptation where

no degradation or very little degradation occurs.

Page 18: Biodegradation overview

Reactions involved in Biodegradation During lag phase induction of enzyme

occurs Duration of acclimatization period varies

from few hrs to days or even weeks depending on biodegradability.

Example: lag phase of 50 days in

pyrazon degradation. III. Recalcitrant/ Xenobiotic: Naturally occurring substances such as lignin

as well as antropogenic.

Page 19: Biodegradation overview

Degradation of petroleum hydrocarbon Aliphatic hydrocarbon belongs to mainly

three groups:Aliphatic

Hydrocarbon

Alkane Alkene Alkyne

Page 20: Biodegradation overview

Alkane biodegradation

Aliphatic hydrocarbons are more saturated compared to aromatic.

Saturation α Biodegradation Branching of the aliphatic chain reduces the

rate of biodegradation. Alkanes are most commonly metabolized by

terminal methyl oxidation. Monooxygenase enzyme plays a key role in

that. O2 MonooxygenaseO atom added to 1° or

2 ° alkane and other atom is reduced to H2O

Page 21: Biodegradation overview

Alkane biodegradation

Reduced NADP that is NADPH2 serve as e-donor and oxidizes alkane aldehyde Fatty acid β- oxidation CO2+ H2O

Sometimes both terminal methyl groups are oxidized results in formation of dicarboxylic acid.

Page 22: Biodegradation overview

Alkane biodegradation

Page 23: Biodegradation overview

Branched Alkane biodegradation

Page 24: Biodegradation overview

Alkene biodegradation

CH3- (CH2)n- CH= CH2

HOOC- (CH2)n-CH= CH2

Sat. end oxidation

CH3- (CH2)n- CH2O= CH2O

Formation of diol

Further oxidation to Carboxylic acid

β- oxidation

Page 25: Biodegradation overview

Degradation of alicyclic hydrocarbon Waxes, Plant Oils, microbial lipids,

Cyclohexane. Hydroxylation of alicyclic alcohol by

Monooxigenase enzyme catalyzed reaction and dehydrogenation by dehydrogenase enzyme leads to formation of ketone.

Further oxidation inserts oxygen into ring and lectone is formed.

Ring cleavageLinearizedAldehydeCarboxylic acid

Page 26: Biodegradation overview

Biodegradation of aromatic hydrocarbons

Most notorious environmental pollutants due to stability.

Example: Polychlorinated Biphenyls (PCBs) Polyaromatic hydrocarbons(PAHs)

The principle reason behind increased resistance to biodegradation is….. Introduction of electronegative groups such as

Chloride, Sulfate, Nitrate. Lowered reactivity of aromatic HC due to

halogen conjugate which decreases interaction with O2

Page 27: Biodegradation overview

Biodegradation of aromatic hydrocarbons

They are oxidized by dioxygenase enzyme which incorporates 2 oxygen atoms leading to formation of CATECHOL.

Dihydroxylated aromatic HC- CATECHOL is cleaved by two ways…

1. Orthocleavage Ring cleavage between two adjacent hydroxyl group by 1,2- Dioxygenase2. Metacleavage Ring cleavage between the carbon atom containing hydroxyl group and adjacent carbon without hydroxyl group by 2,3- Dioxygenase

Page 28: Biodegradation overview

Example: Benzene biodegradation Conversion of benzene to Catechol:

Page 29: Biodegradation overview

Orthocleavage: Benzene biodegradation

Page 30: Biodegradation overview

Metacleavage: Benzene biodegradation

Page 31: Biodegradation overview

Crude Oil biodegradation

Crude oil= aliphatic HC+ alicyclic HC+ aromatic HC

Auto- oxidation in absence of light plays minor role because low temperature of marine environment provide no opportunity for activation.

However, photo- oxidation contribute significantly for self purification of marine environment.

Lab. experiments suggests that, 8 Hrs of effective photoemission may destroy 0.2 metric tons of oil per square Km.

Page 32: Biodegradation overview

Crude Oil biodegradation

More than 100 spp of bacteria, yeast and fungi are capable of oxidizing crude oil.

For ex. Pseudomonas spp, Methanomonas spp, Nocardia spp

Since oil is deficient in some microbial nutrients(especially N, P), so nitrate & phosphate are added to accelerate mineralization.

Optimum temperature: 20- 35ºC Free/ Dissolved oxygen Turbulent condition

Page 33: Biodegradation overview
Page 34: Biodegradation overview
Page 35: Biodegradation overview

Biodegradation of halogenated HC Carbon halogen bond- highly stable and

cleavage of this bond requires substantial energy input.

It is an endothermic reaction. In aliphatic halogenated compound, complete

degradation occurs in two stages- 1. Removal of halogen2. Degradation of organic entity Removal of halogen occurs by two possible

mechanisms….

Page 36: Biodegradation overview

Biodegradation of Halogenated HC 1. Elimination of hydrogen halide:

Direct removal of hydrogen halide between two adjacent carbon atom yields double bond and such reaction occurs rarely.

Page 37: Biodegradation overview

Biodegradation of Halogenated HC 2. Substitution of halogen group by…(a) –H group (Reductive reaction)

(b) -OH group

Page 38: Biodegradation overview

Biodegradation of Halogenated HC S (thio) group:

Most common type of reaction is –OH, which incorporate reactive oxygen group into it.

Page 39: Biodegradation overview

Biodegradation of Halogenated HC Elimination of halide occurs by two possible routes:(a) Elimination of halide after ring cleavage (b) Elimination of halide before ring cleavage

(A) Elimination of halide after ring cleavage: Aerobic degradation of chlorinated aromatic compound usually achieved by a sequence of reaction. HydroxylationCleavage- aromatic ringElimination of cl- from aliphatic intermediate.

Page 40: Biodegradation overview
Page 41: Biodegradation overview

Biodegradation of Halogenated HC The pathway

1. Formation of cl- catechol: The key intermediate in degradation of many chlorinated compounds.

Page 42: Biodegradation overview

Biodegradation of Halogenated HC The pathway

2. Oxidation by Orthocleavage & Metacleavage:

Orthocleavage leads to ring cleavage by 1,2 dioxygenase enzyme followed by elimination of halogen entity. The remaining non halogenated product can be metabolized further.

Metacleavage produces highly toxic intermediate and can’t be taken further by organism involved in biodegradation.

Page 43: Biodegradation overview

Ortho cleavage

Page 44: Biodegradation overview

Meta cleavage

Page 45: Biodegradation overview

Biodegradation of Halogenated HC The pathway

(B) Elimination of halide before ring cleavage:This is not a common pathway. 1. Formation of chlorocatechol:

Page 46: Biodegradation overview

Biodegradation of Halogenated HC The pathway

2. Further oxidation by ortho & Meta cleavage: Ortho cleavage:

Page 47: Biodegradation overview

Biodegradation of Halogenated HC The pathway

Meta cleavage: