1. Investigating the Use of Anaerobic Fermentation onPretreated Biomass to Streamline Bio-fuel ProductionStreamlinedBiofuel ProductionNext Right

2. Contents Hypothesis Controls - Variables Bacteria Studied Methods and Materials Results Acknowledgements 3. HypothesisIf compared with last years study ofenzymatic hydrolysis, single-strainbacterial cellulose hydrolysis will beproven to produce more ethanol;whereas combining two strains ofbacteria in a co-culture will yield thehighest percentage of ethanol. 4. ControlsThe tools, equipment, materials, andprocedures were identical within eachof the three groups studiedThe two different bacteria strains andthe co-culture were the variablesVariables 5. Bacteria StudiedClostridium ThermocellumClostridium ThermolactiumCo-Culture 6. Collect andDry MaterialsGrind, Detoxifyand NeutralizeRecover andMeasureEthanolGraphicalAnalysisBacterial HydrolysisFermentation ofSugarsNoPretreatmentAcidPretreatmentCo-CultureMedia andAutoclaveClostridiumThermolactiumClostridiumThermocellumCorn Stover 7. Glucose and Xylose PercentagesTreated SampleOneTreated SampleTwoUntreatedSample OneUntreatedSample TwoGlucosePercentage48.4 49.2 33.1 33.8XylosePercentage17.3 19.1 16.3 13.5AveragePercentageGlucose48.8 33.5AveragePercentageXylose18.2 14.9 8. Where:mpaper+lignin = Oven dry weight of filter paper and lignin, mgmpaper = Oven dry weight of filter paper, mgmsample = Oven dry weight of sample, mg 9. Klason Lignin ContentTreatedSampleOneTreatedSampleTwoUntreatedSampleOneUntreatedSampleTwoPercentage 17.94 17.62 25.18 24.59AveragePercentage17.78 24.89 10. Basal MediumChemical Formula Required Grams (g)Sodium Chloride NaCl 10.000Magnesium MgCl2.6H2O 0.500Potassium DihydrogenPhosphateKH2PO40.200Ammonium Chloride NH4Cl 0.300Potassium Chloride KCl 0.300Calcium Chloride Hydrate2X with WaterCaCl2 2H2O0.015Sodium Bicarbonate NaHCO3 2.520Resazurin 0.050Yeast extract 4.000L-Cysteine 0.240 11. Results Based on high ethanol content, itwas concluded that the most viablechoice for large-scale productionwas the co-culture Clostridium Thermocellum producedmore ethanol than ClostridiumThermolactium in the single-straintrials 12. HPLC Results Ethanol Content Ethanol AverageBacteriaEthanol, mlethanol per mlof solutionEthanol,% v/vAverageEthanol %(a,b,c)v/vClostridiumThermocellum 1a 0.0720 7.20 7.181b 0.0545 5.451c 0.0890 8.90Control 1d 0.0025 0.25Control 1e*ClostridiumThermolactium 2a 0.0435 4.35 5.082b 0.0410 4.102c 0.0680 6.80Control 2d 0.0012 0.12Control 2e*Co-culture 3a 0.1705 17.05 14.553b 0.1225 12.253c 0.1435 14.35Control 3d 0.0210 2.10Control 3e 0.0180 1.80* The label "*" control samples couldnt give integratable HPLC curves, probably they were toosmall, and were covered by noisy signals. 13. 0246810121416ClostridiumThermocellumClostridiumThermolactiumCo-culturePercentage(v/v)Bacterial Hydrolysis and FermentationEthanol Percentage (v/v) Comparsion 14. Comparison of Enzymatic Hydrolysis (2012) and Bacterial Hydrolysis (2013)Enzymatic Hydrolysis Bacterial Cellulose HydrolysisBiomass(2011-2112)NaOH PretreatmentAverage Ethanol %(a and b)v/v(2011-2012)H2SO4 PretreatmentAverage Ethanol %(a and b)v/v(2012-2013)ClostridiumThermocellumAverage Ethanol %(a,b,c)v/v(2012-2013)ClostridiumThermolactiumAverage Ethanol %(a,b,c)v/v(2012-2013)Co-cultureAverage Ethanol %(a,b,c)v/vCornStover 5.135 8.994 7.18 5.08 14.55Figure A11: Comparison of Ethanol Content 2012 and 2013Comparison of Ethanol Content 2012 and 2013 15. 0246810121416NaOHPretreatmentH2SO4PretreatmentClostridiumThermocellumClostridiumThermolactiumCo-culturePercentage(v/v)Ethanol Percentage (v/v) Comparsion 2012-2013Enzymatic Hydrolysis and YeastFermentation 2012Bacterial Hydrolysis andFermentation 2013 16. AcknowledgmentsI thank Dr. Ulrike Tschirner, from theUniversity of Minnesota, for her generousand steadfast assistance with equipmentand materials, with bacteria, and withinformed counsel.