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Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
Biodegradation
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Biodegradation

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Guest lecture given to the University of Greenwich MSc in in Environmental Science class on 9 February 2004. …

Guest lecture given to the University of Greenwich MSc in in Environmental Science class on 9 February 2004.

N.B. Contact details are out of date.

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  • Until about 1990, it was generally considered that hydrocarbons were essentially immune to anaerobic degradation. Since than, a large number of papers have been written about the degradation of, especially, BTEX compounds. Much of this has been centred around Derek Lovley at the University of Minnesota.
  • If a terminal isomer of LAB were degraded via this pathway, the end result would be either toluene or ethylbenzene, depending on whether the number of carbon atoms in the original alkane chain was odd or even. However, since the pathway requires the addition of molecular oxygen (at two points in each cycle) it will not occur in anaerobic conditions. In any event, the terminal isomers are absent in synthetic LAB.  -oxidation of the isomers found in cable oil may lead to one of a number of structures. This will be dependant on whether the initial chain(s) are odd or even in length, whether both ends of the chain are attacked, and on how close to the phenyl group the relevant enzyme can operate before the charge or physical size of the group interferes too much. However,  -oxidation does require molecular oxygen and so is unlikely to occur in strictly anaerobic conditions.
  • If a terminal isomer of LAB were degraded via this pathway, the end result would be either toluene or ethylbenzene, depending on whether the number of carbon atoms in the original alkane chain was odd or even. However, since the pathway requires the addition of molecular oxygen (at two points in each cycle) it will not occur in anaerobic conditions. In any event, the terminal isomers are absent in synthetic LAB.  -oxidation of the isomers found in cable oil may lead to one of a number of structures. This will be dependant on whether the initial chain(s) are odd or even in length, whether both ends of the chain are attacked, and on how close to the phenyl group the relevant enzyme can operate before the charge or physical size of the group interferes too much. However,  -oxidation does require molecular oxygen and so is unlikely to occur in strictly anaerobic conditions.
  • A number of authors have investigated the degradation of BTEX compounds Grbic-Galic 1991
  • Proposed by Chee-Sanford et al 1996 Note Benzoyl CoA
  • Heider et al 1991 Bemzoyl CoA
  • Heider et al 1999 Benzoyl CoA
  • Benzoate
  • Benzoate, or more specifically, benzoyl-CoA is a central metabolite in anaerobic degradation pathways of aromatic compounds and it’s degradation is fairly well understood. These pathways are proposed from studies of the phototropic bacterium Rhodopseudomonas palustris and the denitrifiers Thauera aromatica (Previously Pseudomona s sp strain K172) and Azoarcus evansii (Previously Pseudomonas sp strain K740) .
  • Aekersberg et al . were the first to show that hexadecane and other long chain alkanes could be degraded to CO 2 by a bacterial strain under sulphate-reducing conditions . There was some evidence that this strain produced membrane lipids with an odd number of carbon atoms when fed alkanes with an even number of carbon atoms. This suggests that the alkane chain undergoes the removal or addition of an odd number of carbon atoms in this organism, which contrasts with the strictly even removal of  -oxidation. This even-to-odd transformation has not been seen in subsequently identified alkane-degrading anaerobes and each species is specific to a limited range of chain lengths, indicating that a range of novel pathways are used.
  • A generalised pathway has been elucidated for the aerobic catabolism of C2-C7 n -alkylbenzenes (i.e. terminal isomers) by Pseudomonas sp. . Alkylbenzenes with an n -alkyl chain of more than 3 carbons are initially attacked by  - or  – oxidation at the methyl terminus. It is possible that both methyl termini are attacked in non-terminal isomers.
  •  -oxidation? No - needs oxygen Conversion to benzoyl CoA? Perhaps - depends on whether alkyl chain degraded Hydrolytic ring cleavage? Most likely (though possibly oxidation under nitrate-reducing conditions) Limited by solubility? May be a plus - low solubility may make for low mobility and low toxicity The most likely route for anaerobic degradation of LABs will probably be an initial attack on the alkyl chain(s) to form an acyl CoA, followed by a hydrolytic ring cleavage.
  • Transcript

    • 1. Biodegradation Dr. Stephen Johnson s.j.johnson@gre.ac.uk
    • 2. Fate of an organic contaminant Volatilization Leaching Sequestration Bioaccumulation Biodegradation
    • 3. Bio – X - ation Biodeterioration – BAD Biodegradation – NEUTRAL Bioremediation – GOOD
    • 4. Microbial biodegradation Aerobic Anaerobic Insitu Ex situ Microbes + contaminants + TEA -> CO2 + H2O + biomass
    • 5. Requirements for life Energy Water Carbon Nitrogen Oxygen? Phosphate Trace elements http://www.abe.iastate.edu/Ae573_ast475/Stoichiom etry_Notes.htm
    • 6. Redox In most cases the contaminant is oxidised (loses electrons). For this to happen, another compound needs to be reduced (gain electrons) to prevent electrons from accumulating. Usually there is a chain of these redox couples with the electrons eventually being taken up by a terminal electron acceptor Oxygen → CO2 Mn(IV) → Mn(III) NO3- → NO2- Fe(III) → Fe(II) SO42- → H2S H → CH4
    • 7. Hydrocarbon degradation Aerobic Nitrate Manganese Fe(III) → Sulphate Methanogenesi Fe(II) s Benzene      Toluene       Ethylbenzene   m-Xylene    p-Xylene   o-Xylene    Alkanes     Alkenes    LAB    PAH    Others     
    • 8. Redox zones Vadose ZoneMethanogenic Water table Sulphate Nitrate Aerobic Saturated Zone Bedrock
    • 9. Aerobic degradation ofn-alkanes β-oxidation Degrades hydrocarbon (fatty acid) chain Removes 2 carbons at a time Ubiquitous pathway BUT needs O2
    • 10. O H2 R C C 115 C C O 112 H H2 2 Fatty acid HS CoA Activation O H2 R C C 12 C C S H2 H2 Acyl CoA CoA Oxidation O H R C C C S H2 H Enoyl CoA CoA H2O Hydration OH O R C C H 42 C C S H2 H2 L-Hydroxyacyl CoA CoA Oxidation O O R C C 56 58 C C S H2 H2 Ketoacyl CoA CoA HS CoA Thiolysis O OR C C 92 S + H3C C 97 S H2 CoA CoA Acyl CoA Acetyl CoA Beta oxidation (Adapted from Stryer 1981)
    • 11. Initial anaerobictransformations of toluene CH3 OH o-Cresol CH3 CH3 CH3 Ring reduction/ Ring cleavage/ Mineralization of Aliphatics Methylcyclohexane Toluene OH p-Cresol CH2OH Benzyl Alcohol
    • 12. Anaerobic mineralizationof toluene O O OH O H2 H C C CH3 C H2C C SCoA H2C C SCoA H H2 H H2C C SCoA H2 Toluene Hydrocinnamoyl-CoA Cinnamoyl-CoA B-hydroxycinnamoyl-CoA O H2O 2e-, 2H+ 2e-, 2H+ SCoA O O CoA O S H2C C SCoA H2 CO2 B-ketocinnamoyl-CoA Benzoyl-CoA 2e-, 2H+ CoASH O SCoA Proposed pathway for anaerobic toluene mineralization - after Chee-Sanford et al 1996
    • 13. Anaerobic degradation oftoluene CoA CoA O O O S O S O O O C C CH3 H H H2C C O H2C C O HC C O H2 H2 H2 Benzylsuccinate Benzylsuccinate- CoA Transferase Benzylsuccinyl-CoA Synthetase Dehydrogenase Toluene Benzyl-succinate Benzylsuccinyl CoA E-Phenylitaconyl-CoA Fumarate 2[H] Succinyl CoA Succinate CoA CoA O S O S O O CoA HO C O C O S H H C C O H C O H2 H2 Phenylitaconyl-CoA 3-Hydroxyacyl-CoA Hydratase Benzoylacetyl-CoA Dehydrogenase Thiolaseolase 2-Carboxymethyl-3-Hydroxy-Phenylpropionyl-CoA Benzoyl-CoA Benzylsuccinyl-CoA H2O 2[H] CoASH Succinyl-CoA Proposed pathway for anaerobic toluene degradation - after Heider et al 1999
    • 14. Anaerobic ethylbenzene degradation CoA S O O O O CoA CH3 HO CH3 O CH3 O CH2 O CH2 O S H H2C C Ethylbenzene 1-Phenylethanol Acetophenone Benzoylacetyl-CoA Dehydrogenase Dehydrogenase Benzoylacetyl-CoA Carboxylase forming enzyme CoA thiolaseEthylbenzene Acetophenone Benzoylacetate Benzoylacetate-CoA Benzoyl-CoA 1-Phenylethanol H2O 2[H] CO2 2[H] CoASH CoASH Acetyl-CoA Proposed pathway for anaerobic ethylbenzene degradation - after Heider et al 1999
    • 15. Anaerobic alkylbenzenedegradation Alkylbenzenes (T,E,X) Key metabolites in Toluene o-Xylene Ethylbenzene m-Xylene p-Xylene degradation COOH COOH COOH COOH COOH H3C COOH COOH COOH COOH COOH All seen in CH3 CH3 laboratory/ground CH3 COOH COOH COOH water CH3 CH3 CH3 Benzoate COOH COOH COOH COOH COOH COOH CH3 CH3 CH3 COOH Elshahed et al 2001
    • 16. Anaerobic benzoatedegradationO OH O SCoA O SCoA O SCoA O SCoA O SCoA B e n z o a t e OH O E3 E4 E5 COO- E6 COO- E2 E7O SCoA O SCoA O SCoA C y c l o h e x - 1 - e n e y d r o x y c y c l o h e x a n e - o C y c l o h e x a n e P i m e l y l - C o A 2 - H 2 - K e t - 2 , 3 - D e d e h y d r o - 1 - c a r b o x y l - C o A - c a r b o x y l - C o A 1 1 - c a r b o x y l - C o A p i m e l y l - C o A HO E1 COO- O SCoA O SCoA E11 E8 O SCoA C y c l o h e x - 1 , 5 - d i e n e 3 - H y d r o x y p i m e l y l -B e n z o y l - C o A 1 - c a r b o x y l - C o A HO HO OH HO O E9 E10 6 - H y d r o x y c y c l o h e x - 2 - e n e - 1 - c a r b o x y l - C o A 2 , 6 - D i h y d r o x y c y c l o h e x a n e - - O x o - 2 - h y d r o x y c y c l o h e x a n e - 6 1 - c a r b o x y l - C o A 1 - c a r b o x y l - C o AE1 -- Benzoyl-CoA reductase E7 -- 3-hydroxyacyl-CoA deyhdrataseE2 -- Cyclohex-1,5-diene -carboxyl-CoA reductase E8 -- Cyclohex-1,5-diene-1-carboxyl-CoA hydrataseE3 -- Cyclohex-1-ene 1-carboxyl-CoA hydratase E9 -- 6-Hydroxycyclohex-2-ene-1-carboxyl-CoA hydrataseE4 -- 2-Hydroxycyclohexane-1-carboxyl-CoA dehydrogenase E10 -- 2,6-Dihydroxycyclohexane-1-carboxyl-CoA dehydrogenaseE5 -- 2-Ketocyclohexane11-carboxyl-CoA hydrolase E11 -- 6-Oxo-2-hydroxycyclohexane-1-carboxyl-CoA hydrolaseE6 -- Pimelyl-CoA dehydrogenase Harwood and Gibson (1997) and Koch et al. (1993)
    • 17. Anaerobic degradation ofn-alkanes  Limited range of chain lengths  No < 6 C to date  Pathways unknown  Specific to organism  May involve addition/removal of odd number of C  Rate of dissolution may limit rate of degradation
    • 18. Aerobic degradation of LAB  If chain > 3 long then starts with β-oxidation of methyl terminus/i  Ring cleavage by oxidation R R R R RCOOH NADH NAD+ H OH C NAD+ NADH OH O + COOH COOH E1 C O2 H OH E2 OH O2 E3 OH E4 OAlkylbenzene Dihydrodiol 2,3-Dihydroxy- Ring fission alkylbenzene product 2-Oxopenta- 4-enoate E1 = Alkylbenzene dioxygenase E2 = cis-alkylbenzene glycol dehydrogenase E3 = 2,3-dihydroxyalkylbenzene 1,2-dioxygenase E4 = ring fission product-hydrolysing enzyme Smith & Ratledge 1989
    • 19. Anaerobic degradation ofLAB β-oxidation? Conversion to benzoyl CoA? Hydrolytic ring cleavage? Limited by rate of dissolution?
    • 20. Generalized breakdown HYDROCARBON (eg BTEX) Anaerobic Aerobic ? Chain degraded by Beta oxidation Convert to e.g.benzoyl CoA Ring cleavage by oxygenases (add O2) Ring cleavage by hydrolysis (add H2O)
    • 21. Organisms Bacteria – bioremediation Fungi – mycoremediation Plants – phytoremediation
    • 22. Bioavailability May not be available to organisms Chemically Physically – Solubility – Sorption
    • 23. Pathways TheUniversity of Minnesota Biocatalysis/Biodegradation Database – http://umbbd.ahc.umn.edu/
    • 24. Bioremediation techniques MNA/MENA Landfarming Bioventing/sparging Windrows Composting Biopile Biofiltration Bioaugmentation Biostimulation Redox/TEA
    • 25. Group 1Not degraded Bitumen Asphalt Metals Inorganic acids Asbestos Complex cyanides
    • 26. Group 2May be degraded in lab Higher MW PAHs PCBs Tars
    • 27. Group 3Demonstrated but not regularly achieved Explosives Pesticides (e.g. lindane, malathion, diuron, mecoprop, paraquat) PCP High MW PAH Branching aliphatics (e.g. hopane) Surfactants (e.g.LAS) MTBE Complex cyanides (?)
    • 28. Group 4Regularly treated aerobically Diesel  Chlorophenols Jet fuel  Organic acids BTEX  Creosote Paraffin  Alcohols Ammonia  Aldehydes Crude oil  Ketones Lubricating oil  Some surfactants Petrol  Some pesticides Phenol  Low MW PAHs
    • 29. Group 5Regularly treated anaerobically Chlorinated solvents
    • 30. Soil factors affecting degradation Organic matter content Microbial activity pH – ionisable compounds – Acid/base catalysed degradation Temperature Soil water content Depth – Faster near surface
    • 31. Phytoremediation Rhizoremediation Transpiration (control flow) Volatilisation Plant metabolism Accumulation Stabilisation/immobilisation

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