Lab - 3 - Biodegradation

1. Lab - 3 - Biodegradation Objectives: Learn the principles of biodegradation process through hands-on experience with aerobic biodegradation processes (Biochemical Oxygen Demand (BOD)) and anoxic biodegradation processes (Denitrification). Also quantify the microorganism biomass in activated sludge biodegradation: Mixed Liquor Volatile Suspended Solids (MLVSS).

2. Lab - 3 - Biodegradation Biodegradation: oxidation of organic matter (CxHyOzNwPv) into mineral components (CO2, H2O, NH3 ammonia, NO3- nitrate) by use of microorganisms Anoxic Biodegradation: Oxidant: nitrate Anaerobic Biodegradation: Oxidant: sulfate Aerobic Biodegradation: Oxidant: oxygen Example: aerobic methanol biodegradation CH3OH +(3/2)O2 => CO2 +2H2O Example: anoxic methanol biodegradation 5CH3OH +6NO3- => 5CO2 +7H2O+3N2+6OH- Important in: Degradation of organic waste Fermentation - Beer!! Decomposition of food. Important in: Removal of nutrients (NO3-) Methane production Food decomposition.

3. Lab - 3 - Biodegradation Biological Oxygen Demand (BOD) Amount of oxygen required to aerobically biodegrade the organic matter (waste) in water Example: Calculate the BOD, expressed as mg of O2/L of water containing 10 mg/L of methanol CH3OH +(3/2)O2 => CO2 +2H2O BOD = 10mg CH3OH/L*(1mmol CH3OH/32mg CH3OH)*(1.5 mole O2/1 mole CH3OH)*32mgO2/ mmolO2 BOD = 15 mg O2/L Real wastewater - very complex: BOD measures the loading of organic matter. Oxygen source for biodegradation: Dissolved Oxygen (DO) in water, for most cases 8 to 9 mg O2/L

4. Lab - 3 - Biodegradation Biological Oxygen Demand (BOD) - Measurement Blank volume, Vb Blank only Sample volume, Vs DOs: dissolved oxygen in sample DOs: dissolved oxygen in blank 300 ml 300 ml Initial DOS,I DOB,I After 7 days Example calculation Vs= 30 ml, Vb= 270 ml DOS,I= 8.5 mg/L, DOB,I= 8.3 mg/L DOS,7= 4.5 mg/L, DOB,I= 7.9 mg/L BOD=((8.5-4.5)-(8.3-7.9)*270/300)*300/30 BOD= 36.4 mg O2/L Final DOS,7 DOB,7 (BOD)7= [(DOS,I-DOS,7)-Vb/300ml(DOB,I-DOB,7)] *300ml/Vs

5. Lab - 3 - Biodegradation Biological Oxygen Demand (BOD) - Measurement For our laboratory 1) Treated wastewater: Vs= 30 ml, Vb= 270 ml of tap water treated with activated carbon 2) Acitvated sludge: Vs = 3ml, Vb= 297 ml of tap water treated with activated carbon 3) Lake water: Vs= 270 ml of lake water Vb=30 ml of treated wastewater 4) Tap water treated with activated carbon Dissolved Oxygen measured with an accumet AP64 series handheld meter, Fisher Scientific brand. Principle of measurement: measure the electric potential of the electrolytic cell for reducing oxygen: O2 + 2H2O + 4e- = 4OH-

6. Lab - 3 - Biodegradation Mixed Liquor Suspended Solids (MLSS) and Volatile Suspended Solids (MLVSS) Total Solids (TS) Includes TDS+TSS Sample Oven, 105ºC overnight 40mm Glass Fiber Filter Filter Cake Vacuum Pump Total Suspended Solids (TSS) Vacuum Filtration Oven, 105ºC, 1h Oven, 550ºC, 20 min Oven, 105ºC overnight Volatile Suspended Solids (Biomass in Activated Sludge) (VSS)=(TSS)-(NVSS) Non-Volatile Suspended Solids (NVSS) Total Dissolved Solids (TDS) :dissolved salts (e.g. NaCl)

7. Lab - 3 - Biodegradation Mixed Liquor Suspended Solids (MLSS) and Volatile Suspended Solids (MLVSS) Sample (one per group): 10 ml (Vs) of activated sludge for denitrification Pre-weigh glass fiber filter (W0) Vacuum filter the sample over a 40 mm glass fiber filter until “cake” consistency - no supernatant water over the filter Dry in air circulating oven for 1h @ 250°C , set in dessicator for 30 minutes and weight the cake+filter (W1) Mixed Liquid Suspended Solid (MLSS) = (W1-W0)/Vs, in mg/L Dry the cake at 550 °C for 20 min, and weigh again= W2 Mixed Liquid Volatile Solids = (W1-W2)/Vs, in mg/L

8. Lab - 3 - Biodegradation Anoxic Biodegradation - Denitrification When nitrate (NO3-) is in water, it is removed by using it as oxidant instead of oxygen to carryout the biodegradation of carbonaceous substrate (e.g. methanol) Example: anaerobic methanol biodegradation 5CH3OH +6NO3- => 5CO2 +7H2O+3N2+6OH- • Process Requirements • Anoxic - Dissolved Oxygen (DO) less than 1 mg O2/L • Facultative Bacteria - Bacteria that can make use of NO3- as oxidant • Pseudomonas, Bacillus, Hyphomicrobium, Moraxella, Paracoccus, etc • Concentration of biomass: MLVSS ~ 1000 mg/L - 5000 mg/L. • Carbon Source: Residual BOD, external source, e.g. methanol • Carbon source loading BOD/NO3--N ~ 3 Example calculation of NO3--N: If 10mg/L NO3--N, then NO3- = 10 mg N * (62 mg NO3-/14 mg N) = 44.3 mg NO3-/L

9. Lab - 3 - Biodegradation Anoxic Biodegradation - Denitrification Setup Denitrification experiments Mixed Liquor: Initial DO (DOI):_____ mg/L Initial NO3--N (NO3-)I :_____ mg/L Note: NO3--N is the concentration of nitrate considering only the atomic weight of nitrogen Required BODR = 3*(NO3-)I mg/L (A) If methanol = BODR/1.5 (B) If ethanol = BODR/2.09 (C) If acetic acid = BODR/1.07 (D) If nothing added, biomass degradation? Initial MLVSS:____mg/L Carbon source:? Volume? Mixed Liquor (300ml) 300 ml Stirrer Measure Nitrate (NO3--N) and dissolved oxygen (DO) every week for three weeks

10. Lab - 3 - Biodegradation Anoxic Biodegradation - Denitrification Project: design a denitrification plant for the NWWTP of Norman, OK Carbon Source (methanol, ethanol,etc) From Aerobic Activated Sludge Clarifier Design the anoxic reactor Clarifier Anoxic reactor [NO3-]=[NO3-]I*exp(-kd*t) NO3- concentration Find the volume of the anoxic reactor, the clarifier, the flow rates and sludge loading. Capital Cost and O&M. Time, hrs