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com185FactorsAffectingtheRateofBiodegradationofPolyaromaticHydrocarbonsShalluSihag1*HardikPathak2andD.
P.
Jaroli21ResearchScholar,DepartmentofBiotechnology,JECRCUniversity,Jaipur,India2AssistantProfessor-IandHeadofDepttofBiotechnology,JECRCUniversity,Jaipur,India*CorrespondingAuthorE-mail:cul.
tanya@gmail.
comABSTRACTInthewholeenvironment,oneofthemostvitalcontaminationsiscausedbythepetrochemicalindustries,products,effluentswhichisreleasedfromthepetrochemicalindustries,roadaccidents,shipaccidentsetc.
Thesepetroleumproductscontainthecarcinogenicandmutageniccompoundswhichareknownaspolyaromatichydrocarbons.
Inthepresentpictureofworldamajorenvironmentalpollutionofsoilandwaterisduetohydrocarboncontaminationresultingbythepetrochemicalindustries,refineries,humanactivitiesetc.
Thebetterunderstandingofthemechanismsandfactorswhichaffectbiodegradationisofgreatecologicalsignificance,sincethechoiceofbioremediationstrategydependsonit.
ForsuccessfulimplementationofbioremediationtechnologiesoncontaminatedareasmustbedependsuponthecharacteristicsofthecontaminatedsiteandacomplexsystemofmanyfactorsthataffectthepetroleumhydrocarbonsbiodegradationprocessesThemainfactorswhichlimittheoverallbiodegradationratecanbegroupedas:soilcharacteristics,contaminantcharacteristics,bioavailability,microorganism'snumber,catabolismevolutionetc.
Inordertoadoptandimplementsomeeffectivebioremediationstrategyitisextremelyimportanttoconsiderandunderstandthoselimitingfactors.
Thepresentstudyexplainssomefactorswhichcantherateofbiodegradationprocess.
Keywords:Polyaromatichydrocarbons;factors;biodegradation;bioremediationetc.
INTRODUCTIONTheintentofthepresentreviewistopresentabroadandupdatedoverviewofthemicrobialecologyofhydrocarbondegradation,emphasizingbothenvironmentalandbiologicalfactorswhichareinvolvedindeterminingtherateatwhichanextenttowhichhydrocarbonsareremovedfromtheenvironmentbybiodegradation.
Aspectsofbiodegradationofpetroleumandindividualhydrocarbonsinmarine,freshwaterandsoilecosystemsarepresented.
Inthemodernindustrialageoftechnologiesvariousaspectsofhumanlifechanges.
Thequalityoflifeonearthishighlyrelatedwithitsenvironment.
Itisveryimportantforlifetosustainthattheenvironmentshouldremaincleanandhealthy.
Inrecenttimesduetotheincreaseinindustrializationandutilizationofpetroleumrelatedproductsnottoforgetnegligenceusingthem,theenvironmentisgettingdamaged.
Tocleanupespeciallythesubsurfacehydrocarbonconcentrationisaworldwideproblem.
Thedevelopmentofhumansocietyalloverhistoryhasleadtorisingdisruptionofthenaturalequilibriumandtherateofdifferenttypesofpollution.
Theplanetdependsonoil,andtheuseofoilasfuelhasleadtointensiveeconomicdevelopmentworldwide.
Thegreatneedforthispowersourcehasledtothegradualexhaustionofnormaloilcapital.
ontheotherhand,mankindwillwitnesstheresultsofoilutilizationforcenturiesafteritstermination.
Ecologicalpollutionwithpetroleumandpetrochemicalproductshasbeenrecognizedasasignificantandseriousproblem1.
Today,themostcommonenvironmentproblemiscausedduetopetroleumproducts.
Iftheseproductsareexposedtotheenvironmenttheycanbehazardoustothesurroundingsaswellastolifeforms.
Millionsoflitersofpetroleumenterintotheenvironment,frombothnaturalandanthropogenicsourceseveryyear.
Availableonlineatwww.
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2(3):185-202(2014)ISSN:2320–7051Ithasbeenestimatedthatglobalproductionofcrudeoilmorethantwelvemillionmetrictonsannuallyandabout1.
7to8.
8millionmetrictonsofpetroleumhydrocarbonescapesintotheterrestrialandmarineenvironment.
Inthewholeenvironmentthereleaseofhydrocarbonswhetheritisaccidentallyorduetohumanactivitiesisamaincauseofwaterandsoilpollution.
Thesoilcontaminationwithhydrocarbonscausesextensivedamageoflocalsystemsinceaccumulationofpollutantsinanimalsandplanttissuemaycausedeathormutations.
Theseaccidentaloilspillscanevencausedamagetotheseaandshorelineorganisms38.
Theothersourcesofcontaminationduetohydrocarbonsincludeservicestations,garages,scrapyard,wastetreatmentplantsandsawmillsetc.
Petroleumisdefinedasanymixtureofnaturalgas,condensateandcrudeoil.
Crudeoilcanconsistofthousandsofindividualcompoundswithhydrocarbonsrepresentingfrom50to98percentofthetotalweightofcrudeoil.
Whenpetroleumcompoundssuchascrudeoilarereleasedintotheenvironment,thecompoundsundergophysical,chemical,andbiologicalchangescollectivelyreferredtoasweathering.
Thedegreetowhichvarioustypesofpetroleumhydrocarbonsdegradeunderthesechangesdependsonthephysicalandchemicalpropertiesofthehydrocarbons14,31,32,42,53.
1.
1PolyAromaticHydrocarbonsThesitescontaminatedbyautomobileorrelatedwithpetrochemicalcompoundscontainavarietyofaliphatic,aromaticandpolyaromatichydrocarbons(PAHs).
Amongthem,PAHsposemorethreattotheenvironmentandhumanhealth18.
ThePAHsconsistsoftwoormorethantwofusedaromaticringsinlinear,angularandclusterarrangements.
ThePAHsarethepollutantswhicharepresentinair,soilandsediments.
PAHsenterintotheenvironmentfromvarioussourcesandtheirderivativesareresultsofincompletecombustionoforganicmaterialsintheenvironment.
Theyarisefromnaturalcombustionlike;forestfiresandvolcaniceruptions,activitiesrelatedtochemicalandpetro-chemicalindustries,agriculturewastes,sludgewastesetc.
PAHsarewidelyfoundinhighconcentrationsatmanyindustrialsites,particularlythoseassociatedwithpetroleum,gasproductionandwoodpreservingindustries.
Duetotheirtoxicity,carcinogenicityandmutagenecitythesePAHshavegainedmuchconcern50,17.
Morethan100differentPAHsareubiquitouslydistributedinenvironment;manyofthempossessmutagenic,carcinogenic,teratogenicproperties58.
Amongthem,16PAHshavebeenidentifiedasprioritypollutantsandsevenofthemclassifiedasprobablehumancarcinogensbyU.
Senvironmentalprotectionagency&EuropeanUnion35.
InrecentyearsthemajorsourcesofPAHspollutionincludeindustrialproduction,transportation,refuseburning,gasificationandplasticwasteincineration.
ThefateofPAHsinnatureisrelatedtoenvironmentduetotheirtoxic,mutagenicandcarcinogenicproperties(phenanthreneisknowntobeahumanskinphotosensitizerandmildallergen,Benzo(pyrene)iscarcinogenicforhumans).
PAHsmaygetabsorbedtoorganic-richsoils/sedimentsandgetaccumulateinfishandotheraquaticorganisms.
Fromtheretheymightbetransferredtohumansthroughfoodchains.
1.
2BioremediationThereareseveralmethodstotreatthecontaminationcausedbypetroleumandpetroleumproductssuchasphysical,chemicalandbiologicaltreatment.
BiologicaldegradationappearstobethemainprocessresponsiblefortheremovalofPAHsinsoil45,54.
Microorganisms,suchasbacteriaandfungimaytransformPAHstootherlessharmfulorganiccompoundsortoinorganicendproductssuchascarbondioxideandwater15,5.
Thebiologicalmethodslikenaturalattenuation,biostimulation,bioaugmentationandbioremediationareefficientandadequatemethodstocleanupsoilwithpetroleumhydrocarbonsascontaminantsbecausethesemethodsdonotadverselyaffectthesite.
ThebioremediationisoneoftheeconomicalmethodscomparedtotheothermethodslikeincinerationandwashingofthesoiletcBioremediationistheoneoftheusefulandinexpensivemethodtoachievetheoptimumbiodegradationcondition,inwhichbytheuseofmicroorganismshydrocarbonsdegradedwhichisthesourcesofcarbonandenergyforthemicroorganisms.
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2(3):185-202(2014)ISSN:2320–7051Bioremediationhaspotentialtoprovidealowcost,non-intrusive,naturalmethodtorendertoxicsubstancesinsoillessharmfulorharmlessovertime.
Currently,researchisbeingconductedtoimproveandovercomelimitationsthathinderbioremediationofpetroleumhydrocarbons.
Onabroaderscope,muchresearchhasbeenandcontinuestobedevelopedenhanceunderstandingoftheessenceofmicrobialbehaviorasmicrobesinteractwithvarioustoxiccontaminants.
Additionalresearchcontinuestoevaluateconditionsforsuccessfulintroductionofexogenicandgeneticallyengineeredmicrobesintoacontaminatedenvironment.
Thesenewtechniquesarebroughtintocommercialpractice,theimportanceofsoundmethodsforevaluatingbioremediationwillincrease.
Andthatwillhelptocreateabetterandcleanerenvironment.
1.
3FactorsThecleaningupofpetroleumhydrocarbonsinthesoilenvironmentisarealworldproblem.
Betterunderstandingofthemechanismsandfactorswhichaffectbiodegradationisofgreatecologicalsignificance,sincethechoiceofbioremediationstrategydependsonit.
Microbialdegradationprocessesaidtheeliminationofspilledoilfromtheenvironment,togetherwithvariousphysicalandchemicalmethods.
Thisispossiblebecausemicroorganismshaveenzymesystemstodegradeandutilizedifferenthydrocarbonsasasourceofcarbonandenergy.
Eveniftheoptimalconditionsformicrobialdegradationareprovided,theextentofhydrocarbonremovalisstronglyaffectedbyitsbioavailabilityandstagesofweathering.
Asaconsequence,somefractionsofhydrocarbonsremainundegraded.
Thisresidualfractionofhydrocarboninsoilcanrepresentanacceptableendpointforbioremediationif1hydrocarbonbiodegradationistooslowtoallowfurtherbioremediation,inwhichcaseothertechnologiesmustbeapplied;2thoseconcentrationsareunabletoreleasefromthesoilandposeadverseeffectstotheenvironmentandhumanhealth,likethosepresentedinthegivencasestudiesVariousstudieshaveaddressedthesuccessfulapplicationsofremediationofhydrocarboncontaminatedsoilandwater.
ItwascarriedoutintegratedstudyofcontaminationcausedbyhydrocarbonandPAHsanditstreatmentwithbioremediationmethod35.
ByinoculatinganenzymaticmicrobialcompounditwasmeasuredconcentrationofhydrocarbonandPAHs.
Someeco-toxicologicalandmicrobiologicaltestswereperformedandreductionsofpollutantswereobtainedwhiletoxicologicalandphytotoxicitytestsevidencedagreatimprovementofthesoil.
Themaximumbiodegradationoccurredwhenconditionsarefavourableformicroorganisms.
Itisimportanttoknowthecharacteristicsofthecontaminatedsitebeforebeginningthetreatments.
Thebasicinformationsuchasresidualoilconcentration,populationdensityofhydrocarbondegradingmicroorganismsandthebiodegradationpotential,environmentalfactorssuchaspH,temperatureetc.
aresomeofthekeyfactorstobeconsideredforbioremediation12,19,41.
DegradationofPAHsinsituisoftenslow,andresearchoverthelastdecadeshasshownthatthesecompoundsveryoftenarepersistent.
Thispersistencemaybeduetoseveralfactorssuchasnutrients,bioavailabilityofPAHs(sorptiontoparticles),temperature,oxygen,andpresenceofPAH-degradingmicroorganisms.
Afullfactorialdesigncanbeusedtotesteverypossiblecombinationofdifferentenvironmentalfactors.
Successfulimplementationofbioremediationtechnologiesoncontaminatedareasdependsonthecharacteristicsofthecontaminatedsiteandacomplexsystemofmanyfactorsthataffectthepetroleumhydrocarbonsbiodegradationprocesses19.
Themainfactorswhichlimittheoverallbiodegradationratecanbegroupedas:soilcharacteristics,contaminantcharacteristics,bioavailability,microorganism'snumberandcatabolismevolution1.
Inordertoadoptandimplementsomebioremediationstrategyitisextremelyimportanttoconsiderandunderstandthoselimitingfactors.
Fortheeffectivebioremediation,informationregardingthephysicochemicalpropertiesandtheindigenousmicrobialcommunityoftheautomobilecontaminatedsoilareessential52.
Alargepartoftheoilresourcesinourplanetisconstitutedbyheavyoil,i.
e.
,oilreservoirsthathavesufferedbiodegradationatacertainextent.
Therearemanyfactorscontributingtotheoildegradation,includingphysical-chemicalfactors,suchastheenvironmentalpH,organicmattercontent,temperature,andtheoilchemicalcomposition,aswellasbiologicalfactors,suchasthemicrobialdistributionintheenvironment,physiologicalandmetabolic.
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2(3):185-202(2014)ISSN:2320–7051Manypublishedstudieshaveinvestigatedtheefficacyofbioremediationonabenchscaleandunderideallaboratoryconditions,suchasacircum-neutralpHandmesophilictemperatures.
However,itisapparentthatenvironmentalfactorsthatvaryfromsitetosite(suchassoilpH,nutrientavailabilityandthebioavailabilityofthecontaminant)caninfluencetheprocessofbioremediationbyinhibitinggrowthofthepollutant-degradingmicroorganisms.
ThereareavarietyoffactorslikepH,temperature,oxygen,nutrientavailabilityetc.
,whichcanaffecttherateofdegradationofpolyaromatichydrocarbons56,57studiedtheeffectoftemperature,salinityandoilconcentrationonbiodegradationofcrudeoil,thebiosurfactantactivityandnaphthalenedegradationbyanewlyidentifiedstrainBacilluscereus28BNwasinvestigatedthemetaboliccapabilityof15bacterialisolatesisolatedfromoilcontaminatedsitebyusingenrichmentculturetechniquewhichwereabletodegradearomaticandpolyaromaticfractions48,26.
Variousfactors,includingtheadditionalpresenceofcarbonsources,pH,moisturecontentandsizeofinoculums,influencedthedegradation2BecauseofthetoxicityandcarcinogenicityofPAHsitismoreimportanttoreduceitseffect.
BythemanipulationinthefactorspHofthesoilmicrobialmanipulationtherateofdegradationofPolyaromaticHydrocarbonscanbeenhanced36.
However,determinationoftheoptimizationvaluesforthedegradationprocessofabioticfactors[(C/N/P/),thenitrogensource,theironsource,theironconcentration,thepHandthecarbonsource]ofPAHs(naphthalene,phenanthreneandanthracene)werecarriedoutbyisolatingbacterialconsortiumbelongingtoC2PL0544.
Table1.
1:-FactoraffectingmicrobialdegradationLimitingfactorsEffectWeatheringAggregation,spreading,dispersion,adsorption.
WaterpotentialEvaporations,photooxidation.
TemperatureOsmoticandmatrixforces,exclusionofwaterfromhydrophobicaggregatesOxidantInfluenceonevaporationanddegradationrates.
MineralnutrientsO2requiredtoinitiateoxidation,NO3-orSO42-tosustain,PHCbiodegradation.
ReactionN,P,FemaybelimitingLowpHmaybelimitingMicroorganismsPHCdegradersmaybeabsentorlowinnumbersTable1.
2:OptimalconditionsformicrobialgrowthandhydrocarbonbiodegradationParameterMicrobialgrowthHCbiodegradationWaterholdingcapacity25-2840-80pH5.
5-8.
86.
5-8.
0Temperature(oC)10-4520-30Oxygen(air-filledporespace)10%10-40%C:N:P100:10:1(0.
5)100:10:1(0.
5)ContaminantsNottootoxicHC5–10%ofdryweightofsoilHeavymetals75Crespectivelyinspent-mushroomcompostduringthedegradationofPAHs.
Over90%degradationofthecontaminatingPAHswasoccurredatthesetemperatures.
VariousenvironmentalfactorswereinvestigatedinthewatersolublefractionofKuwaitcrudeoil.
ResultswereindicatealmostallPAHsweredegradedat150Candatanoxygenlevelof4ppmontheotherhandat400CmostofPAHsdegradedoptimallyat0ppmoxygenlevel59.
Themicroorganismswhichdegradehydrocarbonsaremostactiveinspecifiedtemperaturerangesthatgoverntheproductionofenzymes.
ThethreeclassesandtheiroptimumtemperaturerangesarePsycrophiles(below200C),mesophiles(150C-450C),andthermophiles(above500C).
Mostoildegradingmicroorganismsareactiveinthemesothermicrangeof200Cto350Candprovideoptimumdegradationratesatthesetemperatures.
Temperaturewilldeterminetoacertainextentthetypesoforganismsthatwillbepresentfordegradation.
Ingeneral,degradationrateswillbeslowerincolderwatersthatinwarmersclimates.
Atlowertemperatures,theoilviscosityincreases,thevolatilityofthelowerchainhydrocarbonsdecreasesandsolubilityincreases,makingtheoilmoretoxicandlessappealingtodegradingmicrobes.
Seawaterrangesfrom-20Cto350C,withover90%oftheoceanshavingatemperaturebelow50C.
Biodegradationhasbeenobservedinthisentiretemperaturerange.
Oneexperimentshowedthatatemperaturedropfrom250Cto50Ccausedatenfolddecreaseinresponserates60.
DegradationhasbeenobservedinArcticiceandfrozentundra,butatnegligiblerates61.
Heatisreleasedduringdegradationprocesses,butafirebyspontaneouscombustionisnotpossible.
Temperaturesdonotexceed710C,thelimitingmaximumtemperatureformicrobialsurvival.
Highertemperatureswouldberequiredformostproductstoignite,butthechanceremainssmallduetothevolatizationofthesemolecules.
Itisanimportantfactortogovernthemetabolicactivityofthedegradingmicroorganismsaswellasphysicalandchemicalnatureofhydrocarbons.
Atthemesophilicandthermophilicrangeoftemperaturesitisbeenfoundthattheenzymeactivityofmicroorganismsincreaseswhichhelpsinincreasingtherateofhydrocarbondegradation.
Thereforemajorlythe300C-400Ctemperatureinthemesophilicrangeorsometimes600Ctemperaturesinthermophilicrangeisused.
Thoughitisbeenalsoobservedthatsomehydrocarbonslike,dieselcanbedegradedatlowertemperaturesi.
e.
between00-100C.
Butatlowtemperaturestheviscosityofoilincreaseswhichsuppressesthespreadingofoilonsurfacewhichmakesthedegradationdifficult.
Moreover,inthemesophilicrangemorevarietyoforganismscanbeavailablefordegradationsomesophilicorthermophilictemperaturesarethebetterchoiceforbioremediation30.
2.
2SoilCharacteristicsSoilcharacteristicsareespeciallyimportantforsuccessfulhydrocarbonbiodegradation,someofthemainlimitingfactorsare:soiltexture,permeability,pH,waterholdingcapacity,soiltemperature,nutrientcontentandoxygencontent.
Soiltextureaffectspermeability,watercontentandthebulkdensityofsoil.
Soilwithlowpermeability(suchasclays)hinderstransportationandthedistributionofwater,nutrientsandoxygen.
Toenablethebioremediationofsuchsoil,itshouldbemixedwithamendmentsorbulkingmaterials(straw,sawdustetc.
),asthebioremediationprocessesrelyonmicrobialactivity,andmicroorganismsrequireoxygeninorganicnutrients,waterandoptimaltemperatureandpHtosupportcellgrowthandsustainbiodegradation62,63.
2.
3pHManysitescontaminatedwithPAHsarenotattheoptimalpHforbioremediation.
Forexample,retiredgasworkssitesoftencontainsignificantquantitiesofdemolitionwastesuchasconcreteandbrick.
LeachingofthismaterialwillincreasethepHofthenativesoiland/ormadegroundofthesite,resultinginlessfavorableconditionsformicrobialmetabolism.
Inaddition,theoxidationandleachingofcoalcreateanacidicenvironmentbythereleaseandoxidationofsulfides.
AsthepHofcontaminatedsitescanoftenbelinkedtothepollutant,theindigenousmicroorganismsatthesiteswillnothavethecapacitytowww.
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ItwasstudiedtheeffectofpHontheabilityofsodiumphosphate(Na3PO4)indegradingpolycyclicaromatichydrocarbons(PAHs)incontaminatedsoil.
ResultsobtainedfromGasChromatography/MassSpectroscopy(GC/MS)analysesindicatedthat23%of2-methylnaphthalenewasdegradedastheoverallhighestpolycyclicaromatichydrocarbonatpH2.
0using2gNa3PO4,whileacenaphthene(1.
7%)wastheleastoveralldegradedpolycyclicaromatichydrocarbonatpH2.
0using4gNa3PO4powder.
AnincreaseinPAHsdegradedtrendwasobservedusing4gNa3PO4asthepHwasincreasedfrom2.
0to4.
0.
Therefore,itiscommonpracticetoadjustthepHatthesesites,forexamplebytheadditionoflime.
(64)PhenanthrenedegradationinliquidculturehasbeeninvestigatedatarangeofpHvalues(pH5.
5–7.
5)withBurkholderiacocovenenas,anorganismisolatedfromapetroleum-contaminatedsoil65.
AlthoughbacterialgrowthwasnotsignificantlyaffectedbythepH,phenanthreneremovalwasonly40%atpH5.
5after16days,whereasatcircum-neutralpHvalues,phenanthreneremovalwas80%.
Sphingomonaspaucimobilis(strainBA2)washowever,moresensitivetothepHofgrowthmedia,withthedegradationofthePAHsphenanthreneandanthracenesignificantlyinhibitedatpH5.
2relativetopH766.
PAHdegradationhasbeenrecordedinanacidicsoil(pH2)contaminatedbycoalspoilbytheindigenousmicroorganisms,withtheconcentrationsofnaphthalene,phenanthreneandanthracenereducedovera28-dayperiod67.
Naphthaleneconcentrationswerereducedby50%insoilsdownstreamofanearbycoalpile,withphenanthreneandanthracenereducedbybetween10and20%.
ItwasfoundthatsomeoftheenvironmentalisolateswereabletobothreducethepHoftheliquidmediafrom9to6.
5within24h,andalsoutilizenaphthaleneasasolesourceofcarbon.
Incontrast,thenaphthalenedegradingmicroorganismsPseudomonasfredrikbergensis(DSM13022)andPseudomonasfluorescens(DSM6506),wereseverelyinhibitedbytheelevatedpH.
Itsuggeststhatinsitumicroorganismsatacontaminatedsitemaybenotonlytolerantofthesiteconditions,butmayhavethepotentialtometabolizePAHsinsub-optimalconditions(inthiscase,highpH).
ThechoiceofpHdependsonthemicroorganismstobeusedforthedegradation.
Fungalstrainsarefoundtocarrybiodegradationatlowertemperatures.
Evenwhenindigenousmicrobialconsortiuminclusiveoffungi,yeastandseveralbacteriaisusedtheyarefoundtosurviveatpH2.
AlthoughtherearecertainbacteriathatarealkaliphilesfoundinalkalinelakesatpH7.
5-10.
Salinitymanyisolatesarecapableofgrowingatsalinitycomparabletoseawater.
Thesignificanthydrocarbondegradationwasobservedwith0.
1-2MNaClwheremaximumwasat0.
4Mi.
e.
almostequivalenttonaturalseawater.
Thoughathighersalinitylevel,thedegradationratewasfoundtodecrease.
2.
4OxygenThoughitisnowwellestablishedthatbioremediationoforganiccontaminantssuchasPAHscanproceedunderbothaerobicandanaerobicconditions,mostworkhastendedtoconcentrateuponthedynamicsofaerobicmetabolismofPAHs.
Thisisinpartduetotheeaseofstudyandcultureofaerobicmicroorganismsrelativetoanaerobicmicroorganisms.
DuringaerobicPAHmetabolism,oxygenisintegraltotheactionofmono-anddioxygenaseenzymesintheinitialoxidationofthearomaticring68.
Waysinwhichtomaintainadequateoxygenlevelsforaerobicmetabolismforinsitutreatmentsarediscussedbelowandincludehydrogenperoxideforsub-surfacecontamination.
Forsurfacecontamination,simplesoiltillingand/ormixing,forexampleusingcompostturners,canaeratecontaminatedmaterialwellenoughtoallowPAHtransformationtoproceed.
Thereisstilldebateastowhetherthebenefitsofanaerobicbioremediationareoutweighedbythenegatives,withtheaerationofcontaminatedanaerobicaquiferssuccessfullyusedtostimulateaerobicmicrobialcommunitiesresultinginsignificantreductionsinPAHconcentrationsingroundwater.
Thishasbeenaccomplishedusinghydrogenperoxide,sodiumnitrate69andperchlorate70.
Inaddition,theaerobicbiodegradationofhydrocarbonshasbeenreportedtobeuptoanorderofmagnitudehigherrelativetoanaerobicbiodegradation71.
However,ithasalsobeenreportedthatratesofanaerobicPAHdegradationunderdenitrifyingconditionswerecomparabletothoseunderaerobicconditions72.
Thoughitappearsthatthefutureofanaerobicbioremediationispromising,thereareseveraldrawbackstothepromotionofanaerobicbioremediation.
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Thishasbeenshownincreosote-contaminatedsediment,wherelimitedbiodegradationofPAHswasseenunderdenitrifying,sulfate-reducingandmethanogenicconditions,59eventhoughtherewasanactivelyrespiringanaerobiccommunitypresentinthesediment.
SimilarresultswerefoundwheninvestigatingthepotentialforPAHdegradationinsedimentsamplesfromSanDiegoBay,California73.
MetabolismofPAHsinthesesedimentsthathadhadlowlevelsofpreviousexposuretoPAHsonlyoccurredafteralonglagperiod,andwaspromotedwhentheywere'spiked'withPAH-contaminatedsedimentsthatcontainedanactivecommunityofPAH-degraders.
ThissuggeststhatthedominantinsitumicrobialcommunitydidnotconsistofPAH-degradingmicroorganismsandthatbioremediationwaslimitedbylownumbersofPAH-degradingmicroorganismsratherthanadverseenvironmentalconditions73.
anotherpotentialdisadvantageofthepromotionofinsituanaerobicbioremediationisthatthegeochemistryofthesubsurfacewillbealteredbytheimpositionofreducingconditions.
Asanenvironmentisdrivenanaerobic,allresidualoxygenisdepleted,andelectronacceptorssuchasnitrate,ferricironandsulfatearereducedduringrespiration74.
Thisresultsinthemobilizationofferrousiron,andthereforethereleaseofphosphateFroiron(III)–phosphatecomplexes.
Bothofthesearetoxictotheenvironment;iron(II)israpidlyoxidizedwhenexposedtooxygen,causinganorangeprecipitateinfreshwaterfrequentlyassociatedwithacidminedrainage75andexcessphosphateinfreshwaterscancauseeutrophication.
Inaddition,thereisoftenaconcomitantincreaseinpH,whichcanresultinthesolubilizationofcarbonatemineralsandthereleaseoftracemetals76.
RespirationwillalsoproducepotentiallypotentgreenhousegasessuchasH2S,CH4andN2O77.
Itisclearthatmoreresearchisneededtofullyunderstandtheimplicationsassociatedwiththepromotionofanaerobicbioremediation.
Thediscoveryofawidediversityofpollutant-transforminganaerobesisasignificantstepforwardinunderstandingtheprocessesinvolvedinbioremediation,andthedesignandapplicationofanaerobicremediationbothinsituandexsitutothecontaminatedsite.
Dissolvedmolecularoxygenisrequiredforrespirationofthemicrobeandisusedthroughoutthesubsequentdegradationpathway.
Requirementsforoxygenuptakearesignificant.
Itusuallytakes3to4mlofdissolvedoxygentooxidize1mlofhydrocarbonstocarbondioxideandwater49.
Thisrelativelyhighdemandisduetothehighconcentrationofhydrogenandcarbonintheoil,butverylowconcentrationsofoxygen.
Surfacewaters,suchasoceans,harbors,andlakes,essentiallyhaveaninfinitesupplyofoxygenduetotheair/waterinterfaceandthewindandwaveaction.
But,oxygenbecomestheratelimitingeffectasdepthincreases.
Atsufficientdepthsandindeepwatersediments,degradationofthehydrocarbonscanturnanaerobicwhentheoxygensupplyisdepleted.
Thus,oilthatisdispersedandsinkstothedeepoceansandiscoveredbysedimentswilltakemuchlongertodegrade.
Oxygenreplenishmentcanbehinderedbylarge,thickpoolsofoilonwatersurfacesduetotheblanketingoftheair/waterinterface.
Thisproblemismostlikelytooccurinmarshes,harbors,andinletsthatrelyontheflushingprocessprovidedbytidalmovements.
Mechanicalremovalisrequiredtoincreasetheairinterfaceboundarynecessaryforoxygenreplenishment.
Forsoilenvironments,theavailabilityofoxygenisdependentonthetypeofsoil,amountofmoisture,andtherateofmicrobialdegradationthathasoccurred.
Oxygenisplentifulonbeachsurfacesandsubsurfaceswherewaveandtidalmechanismscontributetore-aerationandreplenishment.
Oxygencontentwillbehighalsonearjetties,pierstructures,andretainingwallssubjecttocrashingwaves.
Oxygenwillbelimitedinfinegrainsoils,beachfrontswithlittleornotidalmovements,andassoildepthsincrease.
Oxygenhasbeen--showntobetheratelimitingstepofdegradationofhydrocarbonsindeepsoilsandgroundwater.
Toalleviatethese3problems,thesoilcanbemechanicallytilledtoprovideaeration,or,fordeepersoilsandgroundwater,oxygencanbe3suppliedbypumpingitintoaseriesofburiedperforatedpipes,bysoilventing,airsparging,injectionofhydrogenperoxide3andbyaeratingthewatertoprovidethenecessaryamountsofoxygen.
Itisoneofthebasicrequirementsforthebiodegradation.
Butagaintheconcentrationofoxygenusedwillhighlydependonthechoiceofmicroorganismused.
Foraerobicbacteria,stoichiometrically3.
1mg/mlofoxygenisrequiredforthedegradationof1mg/mlhydrocarbonswithouttakingintoconsiderationthetotalmassofbacteria.
Soitmayvarywithincreasingordecreasingmassofbacteria7.
Evenanaerobicbiodegradationhasproveditsimportance.
Manydifferenttypesofbacteriaaretestedandfoundtobewww.
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2.
5NutrientavailabilityInadditiontoareadilydegradablecarbonsource,microorganismsrequiremineralnutrientssuchasnitrogen,phosphateandpotassium(N,PandK)forcellularmetabolismandthereforesuccessfulgrowth.
Incontaminatedsites,whereorganiccarbonlevelsareoftenhighduetothenatureofthepollutant,availablenutrientscanbecomerapidlydepletedduringmicrobialmetabolism61.
Thereforeitiscommonpracticetosupplementcontaminatedlandwithnutrients,generallynitrogenandphosphatestostimulatetheinsitumicrobialcommunityandthereforeenhancebioremediation78,79.
TheamountsofNandPrequiredforoptimalmicrobialgrowthandhencebioremediationhavebeenpreviouslyestimatedfromtheratioofC:N:Pinmicrobialbiomass(between100:15:380and120:10:181.
However,arecentstudyhasshownthatoptimalmicrobialgrowthandcreosotebiodegradationoccurredinsoilwithamuchhigherC:Nratio(25:1)thanthosepredictedfromtheratioinmicrobialbiomass,withlowerC:Nratios(5:1)causingnoenhancementinmicrobialgrowth80.
ThelevelofnutrientsrequiredforPAHtransformationaregenerallythoughttobesimilartothoserequiredforotherorganicpollutantssuchaspetroleumcompounds.
However,littleworkhasbeendoneregardingthemostfavorablenutrientlevelsrequiredfortheoptimaldegradationofPAHs,andfurtherworkinthisareawouldbenefitfuturebioremediationtrials.
Itisworthnotingthatfungiareabletoeffectivelyrecyclenutrients(specificallynitrogen),andthatexcessivelyhighnutrientloadingsmayinfactinhibitmicrobialmetabolism.
Inaddition,thehighmolecularweightPAH-oxidizingligninolyticenzymesofthewhite-rotfungiareproducedundernutrientdeficient(oftenlownitrogen)conditions82.
Itthereforeappearsimperativethatthenutrientstatusofthesiteisestablishedpriortothesupplementationofthesitewithadditionalnutrients.
Eventhoughmicrobialmetabolismmaybetemporarilyincreased,thelong-terminhibitionoffunctionallyimportantorganismsmayresultinthefailureofthebioremediationofhighmolecularweightPAHs(suchasbenzo[a]pyrene).
Nutrientsarerequiredtosupportthebiologicalactivity,andhencebioremediation.
Microorganismscommonlyrequirecarbon,nitrogenandphosphorousforthedegradationofhydrocarbons.
Theamountofvariousnutrientsandratioofparticularly,nutrientslikeC,NandPisquiteconceivableregardingthesuccessofthebioremediationprocess.
Variousstudieshavebeenconductingtoevaluatethefactorswhichenhancetheremediationprocess.
ItwasinvestigatedthedegradationofphenanthrenebynaturalmicroflorapresentinseawatersamplesfromGuayanillabaybytheadditionofKNO3asasourceofinorganicnitrogenwhichwasresulted10foldincreaseinthephenanthrenedegradation54.
Theorganiccarboncontentinhydrocarboncontaminatedsiteisfoundtobeveryhighattributedtoconstantinputofhydrocarbons.
Becauseofthehighcarboncontentofoilandthelowlevelofothernutrientsessentialformicrobialgrowth,therateandextentofdegradationare,ingeneral,limitedbythelowavailabilityofnitrogenandphosphorus.
Consequently,growthofhydrocarbon-degradingbacteriaandhydrocarbondegradationcanbestronglyenhancedbyfertilizationwithinorganicNandP.
InmajorityofthetreatmentstheC:N:Pratioismaintainedas120:10:138,21.
Thetypesandquantitiesofnutrientspresentinthesystemplayamuchmoreimportantroleinlimitingtherateofhydrocarbondegradation.
Manystudieshaveshownthataninadequatesupplyofnutrientsmayresultinaslowerrateofdegradationforhydrocarbons(Roberts,1992).
Thecapacityofthemicroorganismtogrowinagivensystemdependsontheorganism'sabilitytoutilizeanyavailablenutrient.
Aerobicmicroorganismsutilizevarioustypesofnutrientsincludingnitrogen,phosphorus,andtraceamountsofpotassium,calcium,sulfur,magnesium,iron,andmanganese.
Nitrogenandphosphorusarevitalnutrientsespeciallysinceoilcontainsverylittleofeither6.
Thelackofeithernutrientwillretardnaturaldegradationrates.
Seawaterisoftendeficientinthesenutrientsbecausenon-oildegradingmicroorganismsconsumethemaswellastheoilconsumingspecies.
Also,phosphorusprecipitatesascalciumphosphateinthepresenceofseawater48.
Concentrationsofnitrogencompoundsinseawaterrangefrom.
1to1mg/l,andphosphorusrangesfrom0to.
07mg/l,dependingonseasonaltemperatures6.
Tocompensateforthelackofnutrients,fertilizershavebeenapplied.
Amendingthesoilinthismannerimprovesthenutritionalstatusofmicroorganismsandencouragesgrowth.
Theamountrequiredtodegradeacertainvolumeofoilhasnotyetbeenthoroughlyunderstood,thoughthesubjectisbeingresearched20.
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6MicroorganismsNumberandCatabolismEvolutionTheabilityofthesoil'smicrobialcommunitytodegradehydrocarbonsdependsonthemicrobe'snumberanditscatabolicactivity.
Microorganismscanbeisolatedfromalmostallenvironmentalconditions.
Soilmicrofloracontainsnumbersofdifferentmicroorganismsincludingbacteria,algae,fungi,protozoaandactinomycetes,whichhaveadiversecapacityforattackinghydrocarbons.
Themainfactorswhichaffecttherateofmicrobialdecompositionofhydrocarbonsare:theavailabilityofthecontaminantstothemicroorganismsthathavethecatabolicabilitytodegradethem;thenumbersofdegradingmicroorganismspresentinthesoil;theactivityofdegradingmicroorganisms,andthemolecularstructureofthecontaminant42.
Thesoilmicroorganismsnumberisusuallyintherange104to107CFU,forsuccessfulbiodegradationthisnumbershouldnotbelowerthan103pergramofsoil.
Microorganismnumberslowerthan103CFUpergramofsoilindicatethepresenceoftoxicconcentrationsoforganicorinorganiccontaminants27,33.
Theactivityofsoilmicrofloracanbecontrolledbythefactorsdiscussedabove-pH,temperature,nutrients,oxygenetc.
Forsuccessfulbiodegradation,itisalsonecessarythatthemicroorganismscandevelopcatabolicactivity,bythefollowingactivities:inductionofspecificenzymes,developmentofnewmetaboliccapabilitiesthroughgeneticchanges,andselectiveenrichmentoforganismsabletotransformthetargetcontaminant27,33.
Hydrocarbonsarearichsourceofcarbonandenergythatmicrobesneedforgrowth.
Beforethecarbonisavailableforuse,largerhydrocarbonmoleculesmustbebrokendownintosimplermoleculessuitableforusebyothermicrobes.
Morethan70microbialgeneraareknowntocontainorganismsthatcandegradehydrocarbonmolecules.
Hydrocarbonsreleasedintheenvironmentarebiodegradedprimarilybybacteriaandfungi.
Thesemicrobesareubiquitousinsoil,freshwater,andseawaterenvironments.
Ofthemanybacteriafoundtodegradehydrocarbons,themostimportantfoundinmarineandsoilenvironmentsareAchromobacter,Acinetobacter,Alcaligenes,Arthrobacter,Bacillus,Flavobacterium,Nocardia,andPseudomonas20.
Amongthefungilisted,Aureobasidium,Candida,Rhodotorula,andSporobolomycesarecommoninmarineenvironmentsandTrichodermaandMortierellaaremostcommoninsoilenvironments20.
Inmarineenvironments,bacteriaareconsideredtobethepredominanthydrocarbonorganism.
Bothbacteriaandfungicontributetohydrocarbondegradationinsoils,withpercentcontributionsrangingfrom50-50to80-20infavorofbacteria.
Evenlessisknownaboutthecomparativerolesofbacteria,fungi,andyeastsinfreshwater,asaretherolesofprotozoaandalgaeinoveralldegradationrates.
Themicroorganismswhichusehydrocarbonsasafoodsourcecanreadilybefoundinvastquantitiesnearplacessubjectedtooilpollution,suchasnaturaloilseeps,shippinglanes,harbors,oilfields,fuelterminals,andsimilarfacilities.
Relativelyfewhydrocarbonutilizersliveinvirginsoilorinthevastopensea.
Fewtononeresideinpetroleumasitemergesfromthedeepunderground,asshownbythevastamountofoilthatstillremainsundergroundevenaftermillionsofyears.
Thepopulationofmicroorganismswillvaryfromsampletosample,dependingonthelocationatwhichitwastaken.
Oilpollutedharborscancontain1EIOA3toIEI0A6microbes/ml.
Intermsofpercentages,waternotpollutedbyhydrocarbonstypicallyhaveonepercentofthepopulationmadeupofhydrocarbondegradingbacteria,whereasinpollutedareastheycanconstitutetenpercentormoreofthetotalmicrobialpopulation15.
Thuscleanupoflongstandingmilitaryfacilitiessuchasfueldepotsandharborsmaybeeasierthanthatofanisolatedaccidentusingbioremediationtechnologyduetoanincreaseofhydrocarbonutilizingmicroorganisms.
2.
7ConsortiumofMicroorganismsAconsortiummaybedefinedasamixtureofdifferenthydrocarbonsutilizingmicroorganism.
Hydrocarbonsrangeinsizefromsingletomanycarbonmolecules.
Themicroorganismsproduceenzymes(groupsofproteinsthatmediateorpromotethetransformationofthehydrocarbonintoasimplercompound)thatattackthehydrocarbonmolecule.
Somemicroorganismscanproduceenzymeswhicharecapableofattackingalmostanysizeortypeofhydrocarbon;otherscanonlyproduceenzymesthatattackandbreakdownonespecifictypeorsizemolecule.
Oncethehydrocarbonisbroken,otherenzymesmayberequiredtofurtherbreaktheremaininghydrocarbon.
Lackofaspecificenzymetoattacktheremainingmoleculefurthermayprovideabarriertowww.
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Thiscomplexseriesofstepsbywhichdegradationoccursiscalledametabolicpathway.
Nosinglespeciesofmicroorganismiscapableofdegradingthemanydifferenthydrocarboncomponentsinoilproducts,thusmanydifferentenzymesandmetabolicpathwaysarerequiredtodegradethesignificantnumberofcompoundscontainedinpetroleumandrelatedproducts.
Whenapetroleumspilloccurs,certainmicroorganismsinthesystemwillexhibitrapidgrowthduetothevastavailabilityofeasilydegradablehydrocarbons.
Thesefastgrowingspeciesmayhinderotherspeciesbydepletingoxygenornutrientsinthesystem.
Whenthoseeasilydegradablehydrocarbonsaredepleted,themicroorganismsmaylacktheenzymenecessarytodegradetheotherhydrocarbonsavailable,anddieoff.
Othermicroorganismscapableofutilizingtheremaininghydrocarbonswillthenexhibitgrowthandflourishinthesystem.
Thus,thecyclecontinues,asspeciesflourishandrecedeasthehydrocarbonstheyexcelatdegradingbecomeavailable,thenaredepleted.
Microorganismsthatreadilydegradethehydrocarbonsfoundinpetroleumproductscanusuallybefoundnearthesurfacesofsoilandwater.
Themainreasonforthisistheavailabilityofoxygen,moisture,andthefoodsource(thehydrocarbon)nearthesurface.
Althoughsomemicrobesareanaerobic(notgrowinginthepresenceofmolecularoxygen),thevastmajorityofmicrobesthatdegradehydrocarbonsareaerobic(thosethatutilizemolecularoxygen).
2.
8BioavailabilityBioavailabilitycanbedefinedastheeffectofphysicochemicalandmicrobiologicalfactorsontherateandextentofbiodegradation83andisbelievedtobeoneofthemostimportantfactorsinbioremediation.
PAHcompoundshavealowbioavailability,andareclassedashydrophobicorganiccontaminants.
(84)Thesearechemicalswithlowwatersolubilitythatareresistanttobiological,chemicalandphotolyticbreakdown84.
ThelargerthemolecularweightofthePAH,theloweritssolubility,whichinturnreducestheaccessibilityofthePAHformetabolismbythemicrobialcell85,86.
Inaddition,PAHscanundergorapidsorptiontomineralsurfaces(i.
e.
clays)andorganicmatter(i.
e.
humicandfulvicacids)inthesoilmatrix.
ThelongerthatthePAHisincontactwithsoil,themoreirreversiblethesorption,andtheloweristhechemicalandbiologicalextractabilityofthecontaminant87.
Thisphenomenonisknownas'ageing'ofthecontaminant.
Thereforethebioavailabilityofapollutantislinkedtoitspersistenceinagivenenvironment.
ReleaseofPAHsfromthesurfaceofmineralsandorganicmattercanbeachievedbytheuseofsurface-activeagents(alsoknownassurfactantsordetergents).
Thesearecompoundsthatcontainbothahydrophobicandhydrophilicmoiety,thusprovidinga'bridge'betweenthehydrophobicPAHmoleculeandthehydrophilicmicrobialcell.
Somemicroorganismscanproducesurfactants(biosurfactants)thatcanenhancethedesorptionofPAHsfromthesoilmatrix88,89.
Thesearepotentiallymoreeffectivethanusingsyntheticsurfactants,astheyarethoughttobelesstoxictotheinsitumicrobialcommunityanddonotproducemicelles,whichcanencapsulatecontaminantPAHsandpreventmicrobialaccess89.
ThebioavailabilityofPAHsinsoilcanbeassessedusingbothchemicalandbiologicalmethods;thoughitisquestionablewhichtypeoftest(s)ismostrepresentativeofthebioavailabilityofhydrophobicorganiccontaminantsinsoil,asbothapproacheshaveinherentlimitations90.
TherearemanybiologicaltechniquestoassessthebioavailabilityofPAHsinsoil.
Thesecanbebaseduponthemonitoringofbiologicalfunction,suchasmicrobialrespirationrates(mineralization)of14C-labelledcontaminants,thebioluminescenceofmicroorganismssuchasluxmicroorganismsand/orlux-taggedpollutantsthatareincontactwithacontaminatedmaterial91andbyassessingthedegreeofdermaldiffusionandgastrointestinalsorptionofPAHsinearthwormsandhencethebioavailabilityandecotoxicityofPAHsinthesoil74.
Inaddition,bioavailabilitycanbemeasuredbymonitoringchangesintheexpressionofgenesthatcodeforPAHdegradationusingmolecularprobes,andalsobytheextractionofsoilpollutantsusingasimulatedmouthandgutdigestivefluidsuchassaliva,inordertodemonstratetheriskthatthecontaminantwillposeifingested.
(92)Biologicalassaysareoftensupportedbyperformingachemicalassayofthebioavailabilityofthecontaminantinthesoil,whichphysicallyextractsthecontaminantfromwww.
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Organicsolventshavebeentraditionallyusedtoextractorganiccontaminantsduringharshextractionprocesses(suchasSoxhletextraction),althoughthisdoesnotdemonstratethetruebioavailabilityofthecontaminant,butthetotalcontaminantconcentrationinthesoil.
However,amorerepresentativeapproachwasusedbyHatzingerandAlexander,86whoextractedthecontaminantswithmildorganicsolvents(suchasmethanol)torepresentthebioavailableproportioninthesoil.
Asmicroorganismscanmostlyonlyaccessthosecontaminantsthatareintheaqueousphase,water-basedsolventsarealsobeingusedtomoreaccuratelypredictthebioavailablefractionoforganiccontaminationinsoil.
Onesuchcompoundishydroxypropyl-β-cyclodextrin90(HPCD),whichcanencapsulatehydrophobiccontaminants.
Inaddition,HPCDdoesnotappeartoinhibitlux-typemicroorganisms,allowingforacombinedbiologicalandchemicalassessmentofbioavailability.
Thebioavailabilityofhydrocarbonsisalsodependentonphysicalstate,hydrophobicity;sorptionontosoilparticles,volatilizationandsolubilityofhydrocarbonsgreatlyaffectstheextentofbiodegradation.
Eveniftheoptimalconditionsforhydrocarbonbiodegradationareprovidedatthefield,ithasbeenshownthataresidualfractionofhydrocarbonremainsundegraded.
Mainly,afteritsarrivalinthesoil,anorganiccontaminantmaybelostbybiodegradation,leachingorvolatilization,oritmayaccumulatewithinthesoilbiotaorbesequesteredandcomplexwithinthesoil'smineralandorganicmatterfractions.
Therateatwhichhydrocarbon-degradingmicroorganismscanconvertchemicalsdependsontherateoftransfertothecellandtherateofuptakeandmetabolismbythemicroorganisms.
Itiscontrolledbyanumberofphysical-chemicalprocessessuchassorption/desorption,diffusion,anddissolution4,8.
Themasstransferofacontaminantdeterminesmicrobialbioavailability.
Theterm"bioavailability"referstothefractionofchemicalsinsoilthatcanbeutilizedortransformedbylivingorganisms.
Thebioavailabilityofacompoundisdefinedastheratioofmasstransferandsoilbiotaintrinsicactivities.
Mostsoilcontaminantsshowbiphasicbehaviour,wherebyintheinitialphaseofhydrocarbonbiodegradation,therateofremovalishighandremovalisprimarilylimitedbymicrobialdegradationkinetics.
Inthesecondphase,therateofhydrocarbonremovalislowandremovalisgenerallylimitedbyslowdesorption.
Altogether,lessbioavailablefractionofhydrocarboncontaminationisformedbyhydrocarbonswhichdesorbslowlyinthesecondphaseofbioremediation22.
Thebiodegradationofanoil-contaminatedsoilcanalsobeseriouslyaffectedbythecontaminationtime,duetoweatheringprocesses,whichdecreasethebioavailabilityofpollutantstomicroorganisms.
Weatheringreferstotheresultsofbiological,chemicalandphysicalprocessesthatcanaffectthetypeofhydrocarbonsthatremaininasoil22,24,42.
Thoseprocessesenhancethesorptionofhydrophobicorganiccontaminantstothesoilmatrix,decreasingtherateandextentofbiodegradation.
Moreover,aweatheredoil-contaminatedsoilnormallycontainsarecalcitrantfractionofcompoundscomposedbasicallyofhighmolecularweighthydrocarbons,whichcannotbedegradedbyindigenousmicroorganisms22,24,3.
Incontrast,arecentlyoil-contaminatedsoilcontainsahigheramountofsaturatedandaliphaticcompounds,whicharethemostsusceptibletomicrobialdegradation.
However,thepollutantcompoundsinarecentlycontaminatedsoilarepotentiallymoretoxictothenativemicroorganisms,leadingtoalongeradaptationtime(lagphase)beforedegradationofthepollutantandeventoaninhibitionofthebiodegradationprocess22,27,33.
Aswasmentionedabove,sequestrationandweatheringoforganiccontaminantsinthesoilreducesthebioavailabilityoforganiccompoundsandresultsinnon-degradedresiduesinthesoil.
Contaminantsthathavebeenweatheredandsequestratedinsoilarenotavailableforbiodegradationinsoil,eventhoughfreshlyaddedcompoundsarestillbiodegradable1.
Sorptionisamajorfactorpreventingthecompletebioremediationofhydrocarbonsinsoil.
Slowsorptionleadstothehydrocarbonfractionbecomingresistanttodesorptionandincreasesitspersistencewithinthesoilorganicmatrix.
Thefollowinghypotheseshavebeenproposedasaexplanationforweathering:1weatheringresultsinaslowdiffusionofthehydrocarbonfractioninthesolidfractionoftheorganicmatterinthesoil;2thecontaminantslowlydiffusesthroughthesoilandbecomessorbedandtrappedinthesoilnano-andmicropores42,46.
2.
9ContaminantCharacteristicsPetroleumhydrocarbonscontainacomplexmixtureofcompounds;allthecomponentsofpetroleumdonotdegradeatthesamerate.
Theratebywhichmicroorganismsdegradehydrocarbonsdependsuponwww.
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Petroleumhydrocarbonscanbecategorizedintofourfractions:saturatesaromatics,resinsandasphaltene.
Thevariouspetroleumfractions,n-alkanesofintermediatelength(C10-C25)arethepreferredsubstratesformicroorganismsandtendtobethemostreadilydegradable,whereasshorterchaincompoundsarerathermoretoxic.
Longerchainalkanes(C25-C40)arehydrophobicsolidsandconsequentlyaredifficulttodegradeduetotheirpoorwatersolubilityandbioavailability,andbranchedchainalkanesandcycloalkanesarealsodegradedmoreslowlythanthecorrespondingnormalalkanes.
Highlycondensedaromaticandcycloparaffinicstructures,tars,bitumenandasphalticmaterials,havethehighestboilingpointsandexhibitthegreatestresistancetobiodegradation.
Ithasbeensuggestedthattheresidualmaterialfromoildegradationisanalogousto,andcanevenberegardedas,humicmaterial3,22,4,45.
Therateofmicrobialuptakeandbiodegradationofhydrocarbonsisdependentonthesolventsolubilityofthehydrocarbons.
Hydrocarbonsthatarehighlysolubleinwatertheirdegradationratesarealmostproportionaltotheconcentrationbutitdiffersforlessaqueoushydrocarbons.
Itneedstobeconsideredthathighsolubilitymaybecomedetrimentaltothedegradingorganisms.
Usuallyoilloadinggreaterthan5%leadstodecreaseinthemicrobialactivity10,12.
InadditiontothishighconcentrationmaydisturbC:N:Pratioandcreateoxygenlimitations.
2.
10Toxicityofend-productsTheprincipleofbioremediationistoremoveordetoxifyacontaminantfromagivenenvironmentusingmicroorganisms.
Mostcommercialbioremediationtrialstendtomonitorthesuccessofthetreatmentbythedegreeofremovaloftheparentcontaminantanddonotconsiderthepossibilityofthebiologicalproductionofmoretoxicbreakdownmetabolites.
However,itisimportanttoensurethatthecontaminatedmaterialissuitablydetoxifiedattheendofthetreatment93,94.
ArecentstudyusingabioreactortotreatPAHcontaminatedgasworkssoilmonitoredboththeremovalofPAHsandtheaccumulationofoxy-PAHs,suchasPAH-ketones,quinonesandcoumarins94.
ThesecompoundsareformedduringthemicrobialmetabolismofPAHsandcanalsobeformedfromchemicaloxidationandphototransformationofPAHs95.
Suchtransformationproductscanbeequallytoxic,ifnotmoretoxic,tohumanhealthwhencomparedwiththeparentPAH,60withmanyoftheoxy-PAHsformedduringthetreatmentofPAHcontaminatedsoilsmorepersistentthantheparentcompounds94.
Inthisstudy,Lundstedtandcolleaguesinvestigatedthatalthoughtherewerenonewoxy-PAHsformedduringthebioremediationofanagedgasworkssoil,theconcentrationsof1-acenaphthenoneand4-oxapyrene-5-oneincreasedinthesoilby30%and60%respectivelyover30daysofbioslurrytreatment.
Inaddition,theyshowedthatsomeoxy-PAHsactuallyincreasedinconcentrationduringtreatment,andweresubsequentlymorepersistenttomicrobialdegradationthantheircorrespondingparentPAHcompound.
Asoxy-PAHsaremoretoxicthantheparentPAHs,thisstudyhighlightstheimportanceofmonitoringthemetabolitesofbioremediation,specificallyfortoxicdead-endproducts,andassessingthetoxicityofthematerialbothbeforeandaftertreatment.
However,itisimportanttounderstandtherelevanceofecotoxicityteststotheoveralltoxicityoftheremediatedland.
Manyofthesetestsmonitorfor'acute'toxicityofcompounds(viaorganismdeath),whereasitwouldbemorerepresentative,particularlywhenassessingthecarcinogenicandmutagenicPAHs,toconsiderthe'chronic'toxicityofthesesoils,suchasmonitoringfororganismDNAdamageandtheoccurrenceofDNAadducts60.
2.
11MoistureMoistureisrequiredforallbiologicalprocessestohelptransportnutrients,foods,andwasteproductsinandoutofthemicroorganisms.
Foroceans,lakes,andothersurfacewaterenvironments,thisposesnoimmediateproblems.
Forsoilenvironments,somemoisturemustbepresentfordegradationtooccur.
Toomuchwatercanimpedethereaerationofthesoil,andtheprocessmayturnanaerobic.
Theoptimumratioofmoisturewilldependontheclimateandsoiltype.
Ratiosrangefrom30-90%inonestudyand12-32%inothersandtheaerobicbiodegradationofhydrocarbonsinsoilsisgreatestinrangesof50-70%ofthesoilwaterholdingcapacity20,36whereaswavesandtidalactionsareusefulinsupplyingaeratedseawatertobeachesandmarshes,rainfallisusefulinthebiodegradationofinlandsoilsbysupplyingmoistureandusefuldissolvedoxygentothemicrobes.
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12OrganicmatterThepresenceofnaturallyoccurringorganicmattercanhavedifferentaffectsonsoils,suchaswaterretention,soiltemperature,andtheabilityofmicroorganismstodegradepollutants36.
Theroleitplaysinsorptionprocessescanaffecttheavailabilityofnutrientsforthemicroorganisms.
Contaminantstargetedforbiodegradationcanberesistantofalterenzymaticattackbysorptionontosoilparticles.
Theboundmaterialbecomesunavailablefortheattack.
Additionoforganicmaterialcanslowthenaturalrateofbiodegradationduetoincreasedcompetition,butlongtermitcanincreaseinfiltrationandpermeabilityandporosity15.
Theadditionofthematerialcanbehelpfulinlowmoistureretainingsoilenvironments.
2.
13OilsurfaceandconcentrationSincethemajorityofthebiodegradationofhydrocarbonsoccursatorneartheair/waterinterfaceinwaterenvironmentsandtheair/soilinterfaceinsoilenvironments,theamountofoilsurfaceareaexposedattheseinterfaceswillaffecttherateofdegradation.
Thegreatertheareaexposed,thefasterstheproductwilldegrade.
Concentrationofthehydrocarbonisrelatedtooilsurfacearea.
Thickrafts,blankets,orpoolsofoilorotherpetroleumproductsconstituteahighconcentration/lowsurfaceareasituation.
Theavailablesitesthatcanbeattackedarereduced.
Theoilactsasablanket,hinderingthereplenishmentofnutrientsoroxygentothemicrobes.
Thus,athighconcentrationsthosecompoundsmostreadilydegradedwillbeattacked,leavingthemoreresistantcomponentsbehind.
Theseinturncombinetoformevenmoreresistantcompounds,suchastarballsthathavelimitedmoisturecontactandsurfacearea.
Concentrationsofhydrocarbonsintherangeof1to100ug/mlofwateror1to100ug/gofsoil(dryweight)arenotgenerallyconsideredtoxictocommonbacteriaorfungi36.
Thus,dispersantsandemulsifiersusedatseaonheavyconcentrationsofoilincreasetheoilsabilitytospreadandvolatilize,whichinturnallowsforanoverallincreaseintherateofdegradation,pendingtherearenoadversetoxiceffectsofthetypeofdispersantused.
Othermeansofremovingheavyconcentrationsofoilandproductsshouldfirstbeemployedpriortoremediation,suchasusingskimmers,vacuumtrucks,adsorbents,andearthmovingequipment.
2.
14SalinityMicroorganismsaretypicallywelladaptedtocopewiththewiderangeofsalinitiescommontotheworld'socean32.
Thereislittleevidencetosuggesttheyareaffectedbyotherthanhypersalineenvironments,suchassaltwaterfromoilwells.
Estuariesmaypresentaspecialcasebecausesalinityvalueswillvaryinlevelsascomparedtolevelsintheocean49.
Thus,ifmicrobesaretobeaddedtotheenvironments,itmustbeknowniftheyarecompatiblewiththesalinelevelspresentinthesystem.
CONCLUSIONInthepresentscenariomajorenvironmentalpollutionofsoilandwaterisduetohydrocarboncontaminationresultingbythepetrochemicalindustriesactivities.
Itcancausebyaccidentalliberationofpetroleumindustriesdischargeintheenvironmentoritcanalsocausebyhumanactivity.
Hydrocarboncompoundsareknownforitscarcinogenicandneurotoxicbehavior.
Manypublishedstudieshaveinvestigatedtheefficacyofbioremediationonabenchscaleandunderideallaboratoryconditions,suchasacircum-neutralpHandmesophilictemperatures.
However,itisapparentthatenvironmentalfactorsthatvaryfromsitetosite(suchassoilpH,nutrientavailabilityandthebioavailabilityofthecontaminantetc)caninfluencetheprocessofbioremediationbyinhibitinggrowthofthepollutant-degradingmicroorganisms.
Thesefactorsplaysignificantroleinthebiodegradationprocessofpolyaromatichydrocarbonandcontaminatedsoils,suchasphysicalconditions,nutrition,theratiosofvariousstructuralhydrocarbonspresent,thebioavailabilityofthesubstrateandthediversityofthebacterialcommunitiesinvolved,pH,waterholdingcapacityofthesoil,microorganisms.
Therearesomestandardconditionsonwhichthesefactorsplaytheirrolelikewaterholdingcapacityofthesoilforthemicrobialgrowthis25-28andforhydrocarbondegradationis40-80.
TherangeofpHisfrom5.
5-8.
8formicrobialgrowthandforhydrocarbondegradationis6.
5-8.
0.
Thefavorableconditionsoftemperatureforthegrowthofmicroorganismsvaryfrom10-450Candforthedegradationofcontaminantsarefrom20-300C.
Therefore,thisminireviewtouchesuponvariousaspectsofenvironmentalfactorswhichaffectsthebiodegradationofpolyaromatichydrocarbons.
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Biosci.
2(3):185-202(2014)ISSN:2320–7051AcknowledgementsIwishtoexpressmysinceregratitudetoDept.
ofBiotechnologyofJECRCUniversityforgivingmeanopportunityfordoingmyPh.
D.
IwouldalsoliketothankDr.
WidhiDubey(DirectorofSchoolofSciences,JECRCUniversity)forhermanualsupport.
AlsoverybigthankstoDr.
S.
K.
Sharma(Ph.
DCo-ordinator,JECRCuniversity)forhisalltechnicalsupportandguidancethroughoutthestudy.
Also,Iwouldliketothankmyfamilyandfriendsfortheirmanualsupport,strength,andhelpthroughoutthestudy.
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