The purpose of this experiment is to perform a Biochemical Oxygen Demand test on primary clarifier effluent from a wastewater treatment plant to determine a BOD versus time curve. This curve can then be used to determine the Ultimate BOD of the wastewater sample and the rate constant for its decay.

1. 1
Laboratory Report
Experiment # 6
Biochemical Oxygen Demand
Group #1
Jonathan Damora
Tuesday March 11, 2014

2. 2
Purpose
The purpose of thisexperimentisto performa Biochemical OxygenDemandtestonprimaryclarifier
effluentfromawastewatertreatmentplanttodetermine aBODversustime curve. Thiscurve can then
be usedto determine the UltimateBODof the wastewatersample andthe rate constantfor itsdecay.
Introduction
Biochemical OxygenDemand(BOD) isarepresentationof dissolvedoxygencontentinrelationtothe
speedatwhichbacteriaaerobicallydecompose,oxidize,organicmatterwithinasample. Thusitisa
methodof definingthe strengthof wastewater,whichitself dependsonthe level of easily
biodegradableorganicmatterwithinagivensample. If there isdissolvedoxygenpresentandorganic
matterthat can be oxidizedbybacteria,the organicmatterwill be decomposedbythe bacteria
aerobicallyuntil the oxygenislowenoughthat anaerobicbacteriabecomesdominant. BODtestingwas
developedover100 yearsago, withone of the earlieststandardizationsof BODtestingonsewage
comingfromthe RoyalCommission on SewageDisposalin1912, whichstatestheirregulationsonBOD
of effluentat65 degreesFahrenheit. BODtestinghassince beenstandardizedat20 o
C.
The testingof BOD is importantthroughoutmanyfields,asBODis listedasa conventionalpollutantin
the CleanWater Act,passedin1972. It is usedto measure the effectivenessof awastewatertreatment
plant, the pollutionalstrengthof industrial wastewater,andiscommonlyusedasa measure of the
strengthof any freshwatersample orpurificationstrengthof abodyof water. The UnitedStatesuses
BOD to manage the qualityof secondarytreatment,whichis expectedtoremove 85% of BOD measured
inthe sewage andproduce effluentthathasa 30-day average BOD below 30 mg/L anda 7 day average
BOD below45 mg/L. Industrial pre-treatmentof wastewaterusesBODtoboth determine the level and
type of treatment needed. If the BOD to COD ratiois greaterthan.4 or .5 thenbiological treatmentof
the wastewaterispossible,aslongasthere are nobiological toxinspresent. The industrial BODtesting

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may require the additionof seedbacteria,asthere maybe no livingbacteriapresentdependingonthe
prioruse of the water.
BOD testingspecificallymeasuresthe dissolvedoxygenconcentrationatdifferentpointstodetermine
the rate at whichoxygenisused. The metabolismof organiccompoundsbybacteriacan be represented
by the belowequation,withthe bacteriaactingasthe host forthe oxidationof organiccompoundsto
CO2,water,and ammonia. Inthiscase the organiccompoundisoxidizedbyoxygen.
Aerobicdecompositionisnotthe onlysource of oxygendemandwithinasample,asthere are
chemoautotrophicbacterianitrifyingammonia,ammonium, andnitrite intonitrate. Thissplitsa
measurementof BODintocarbonaceousBOD (cBOD) and nitrogenousBOD(nBOD),unlessanitrification
inhibitorisused,suchasthe 2-chloro– 6- (trichloromethyl) pyridine usedinthisexperiment. Usually
for a BOD5 testdone at 20 o
C the small numberof nitrifyingbacteriapresentdonotaddsignificantlyto
the oxygendemand,butourexperiment goesto9 days whichallowsthe populationof nitrifying
bacteriato grow toan appreciable level. The demanddue tonitrifyingbacteriaisnotconsideredas
importantindeterminingpollutionstrengthascBODsince nBOD doesnotinvolve metabolismof organic
matter. Otherinterferencesincludebio-inhibitionfromtoxins,non-bioassimilable organicmatter,and
otherformsof oxygendemand.
The valuesobtainedbyaBOD test are usedto determine the ultimate BODof the wastewatersample
and the rate constantfor itsdecay. These factorsarise fromempirical studiesof the decayof
biodegradablematterinwastewater,whichshow thatitusuallyfollowsfirstorderkinetics. Thismeans
specificallythatthe rate at whichorganicmatter isoxidizeddependsonthe amountof organicmatter

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present. Thusthe BOD at a specifictime,t,canbe calculatedfrom
the above equationonce ultimate BODandthe rate constant,k, are determined.
Procedure
To performour BOD testwe made 48 identical 300-mLBOD bottlescontaining8mL of settled
wastewater(primaryclarifiereffluent) and 292 mL of a solutioncontaining:aerateddistilledwater,
ferricchloride,magnesiumsulfate,calciumchloride,aphosphate buffer,andthe nitrificationinhibitor
(2-chloro– 6- (trichloromethyl) pyridine). The otherchemicalsare addedtoprovide nutrientsand
bufferpHin orderto facilitate bacterial growth. These bottleswere thenplacedintothe BODincubator
at 20 o
C exceptfor4 of the bottleswhichare usedtodetermine the initial DOconcentrationusingthe
Winklertitrationtechnique. The DOconcentrationisdeterminedeachdayfor9 days,exceptthe 6th
and
8th
day,witheach day’stitrationbeingreplicated using6separate BOD bottleswiththe DObeing
averagedforthat day.
The DO concentrationisdeterminedusingthe WinklerTitrationtechniqueoutlinedhere. Firstwe
added1 mL of manganese (II) sulfate andalkali-iodide-azide solutionrespectivelytoourBOD bottle and
thoroughlymixedit. A brown precipitateisformedinthe presence of dissolvedoxygen,ora white
precipitate of Mn(OH)2 isformedinthe absence of DO. If there isno DO,thenthe processis complete
since the purpose isto testthe concentrationof DO. If there isoxygenanda brownmanganese
precipitate forms,add1mLof sulfuricacidandmix thoroughly. The brownprecipitate dissolvesandthe
Iodide ion(I-
) isconvertedtoiodine. Titrationisthenperformedusingasodiumthiosulfate solution and
a starch indicator,withthe DO concentrationequal tothe volume of sodiumthiosulfateused. Asstated
above,these stepstodetermineDOconcentrationwere replicatedupto6 timeseachday and average
valuesare presented.

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Figure 1. BOD-Time Curve
Results
0.00
50.00
100.00
150.00
200.00
250.00
300.00
0 1 2 3 4 5 6 7 8 9 10
BOD(mg/L)
Time (days)
BOD-Time Curve
Figure 2. Graphical Determination of k
y = 0.0151x + 0.2065
R² = 0.9691
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0 1 2 3 4 5 6 7 8 9 10
(t/BOD)^(1/3)
Time (days)
Graphical Determination of Rate Constant, k

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Table 1. BOD Results
Time
(day)
Average
DO (mg/L)
BOD
(mg/L) (t/BOD)^(1/3)
Ultimate
BOD
0 7.70 0.00
1 4.80 108.75 0.21 305.6228
2 3.80 146.25 0.24 249.9804
3 3.00 176.25 0.26 240.5489
4 2.30 202.50 0.27 244.6204
5 2.10 210.00 0.29 236.1985
7 1.90 217.50 0.32 227.9936
9 1.20 243.75 0.33 248.4959
The interceptof the bestfitline forFigure 2 is 0.2065 andthe slope is0.0151, thus
k=2.61*(.0151/.2065)=0.191. Thisrate constantcan be pluggedintothisequation.
Alongwiththe k value determinedabove Iwill plug tandBOD at each time. The resultsare tabulatedin
Table 1, withthe average ultimate BODequal to250.5 mg/L.
The BOD inthe above table wascalculatedusingthisequation, . Where D1 is
the initial DOconcentration,D2 isthe DO at time t, P isequal to the decimal fractionof wastewater(8ml
wastewater/300ml BOD bottle volume),andBODisthe calculatedBODat time t.
Discussion
The resultsfitwhatwas expectedfromasample containingorganicpollutants.The BOD-time curve
displaysthe distinctiveplateauasthe microbial populationshiftstowardspredominantlyprotozoa
predationandthenincreasesagainasthe deadprotozoacellsare decomposedbymore bacteria.The k
value of 0.191 isaroundthe expectedvalue,althoughslightlyhigher,whichmeansthatdecomposition

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occurredrapidlyandthere was notany biological growthinhibitorspresent. The ultimate BODresultof
250.5 mg/L ispossiblymisleadingsincethere ismuchdebate overthe use of firstorderchemical kinetics
on BOD. WhenusingBOD = BODo (1-e-kt
) the ultimate bodvalue ismuchhigher,around450-550 mg/L.
Andwhenusingthe methodgiveninSnoeyinkandJenkins’ WaterChemistry, the Ultimate BODis594.
The highervaluesseemmore likelysince the BOD-timecurve showsthatthe BOD isclimbingagain at
day 9. There is mostlikelynonitrificationinterference due tothe nitrificationinhibitoradded. Itis
possible thatotherinterferencesoccurred,suchasmicrobial inhibition,althoughunlikelysince the
resultwasfairlyhigh.
DiscussionQuestions
1. What are the applicationsof BODtestdata? DiscussBOD testlimitationintermsof the
biodegradabilityof wastes.
BOD testinggivesdatathatshowsthe relative organicstrengthof thatwatersample as well asthe
decayrate of said organicmatter.BOD testingisapplicable throughoutindustrial andpublicwastewater
treatment. The US Governmenthasevenputregulationsonthe General Pollutant,BOD,withinthe
effluentdischargedfromwastewatertreatmentplants. Forexample,testingthe BODof wastewater
priorto treatmentandthenagain aftertreatmentwill tellyouthe efficacyof yourtreatmentprocess,in
termsof organicpollutionalstrength. BODtestingattemptstofocusspecificallyonoxidationof organic
compoundsusingoxygen,butthere are manydifferentformsof bacteriathatuse differentoxidizing
agents. For example,thereare speciesof bacteriathatoxidize ferricironintoferrousiron. The BOD
resultscannotshowyouexactlywhatcontributedtothe DO demand,thusitisdifficulttodetermine
whetherthere isanyinterferencesthatcontributedtoyourdata otherthanthe biodegradationof
organicwaste.

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2. What purposesare servedbyferricchloride,magnesiumsulfate,phosphoricacid,ammonium
chloride,andcalciumchloride?
FerricChloride isusedasa coagulantto settle suspendedsolids,aswell asbeinganimportantmineralin
aerobicprocesses. MagnesiumSulfate isacommonformof the mineral Magnesium. Phosphoricacidis
usedas a pH buffertoensure itstays constant,as well asa wayto add phosphate tothe solutionwhich
allowsanyferricoxide withinthe solutiontobe precipitatedout asferricphosphate. Ammonium
Chloride isusedtobufferthe pHof the solution. CalciumChloride addsbioavailable calciumionsinto
the solution,whichisanecessarymineral formanyformsof life.
3. How dothe BOD resultsrelate toCODdata from previousLab?
We couldcompare the rationof BOD/CODto determine the levelof oxygendemandfrommetabolism
of organicproducts,whichwouldgive usanindicationwhetherbiological treatmentwouldbe efficient
to use on the wastewater. The wastewaterin the previouslabgave aCOD value of approximately415
mg/L,and thisexperimentdeterminedthatthe BODof the wastewaterwas250.5, thus the rationof
BOD to COD is.6, if the twosamplesare fromthe same source. Thismeansthat biological treatment
wouldstill be effective onthiswater. The CODtest wasmuch lesstime consumingbutgave less
detaileddata.
4. List the majorfactors influencingthe rate of biologicaloxidationinthe BODtest.
The major factorsfor BOD testingare of course firstof all the testparameters,meaningthe organic
matterpresentandthe dissolvedoxygenconcentration. Thenthe largestfactorsonBOD resultsare
nitrifyingbacteria,since theywill increase the demandforDO,andtoxiccompoundsorresidual chlorine
fromtreatment,whichwill inhibitmicrobiallifewithinthe sample thusloweringBODresultsincorrectly.
Otherfactors include the pHof the sample,whicheffectsthe equilibriumof manyreactionsandchanges

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speciesconcentration,temperature of the sample and the presenceof certainnon-assimilable organic
material suchas woodpulp,whichwill increasedemandforDO.
Conclusion
BOD testingisa useful waytodetermine the levelof biological activityandthe concentrationof organic
matterthat is useful tomicroorganisms. The longtestingtime detractsfromthe attractionof thistest
overCOD, since itfocusesspecificallyonthe organicmatterthat bacteriacan decompose. Despiteits
limitationsitisclearthat BOD testingisessentialasitallowsusto quantifyorganicpollutionwithina
watersample,withoutexpensive chromatographyorothermethodsusedtodeterminecomposition.

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Works Cited
Snoeyink, Vernon L., and David Jenkins. Water Chemistry. New York: Wiley, 1980. Print.
"BOD." Wikipedia. Wikimedia Foundation, 10 Mar. 2014. Web. 11 Mar. 2014.
"Winkler Titration Technique." Wikipedia. Wikimedia Foundation, 10 Mar. 2014. Web. 11 Mar.
2014.