By: Zachary Bryan Scott Department of Civil and Environmental Engineering University of California, Irvine

Motivation Wastewater Treatment: Aerobic Biological Treatment of Wastewater Cleans wastewaters Generates Biomass (0.4g VSS/ g COD) Anaerobic Biological Treatment Cleans wastewaters – Many Types Reduces biomass (0.05 g VSS/ g COD) Can produce net energy Consider heating, transport, 4 ft 3 CH 4 / lb VS as manure 2

Potential Uses of Process Models: 3 Reactor Assessment/Optimization Design Process Modification

Do Current Process Models Make Predictions of Sufficient Accuracy? 4 Modify Reactor Conditions: pH, T, Feed Type, Feed interval, Mixing Intensity, Nutrients Current Industrial Use of Models is very limited Pilot testing still dominant = higher costs = more time

5 Modified figure, Gujer and Zhender, 1979 Anaerobic Process:

Three Critical Measurements: Methane Production Rates- Flowcell Acetate Consumption Rates-GC Aceticlastic Methanogen Ribosomal DNA Copy # - qPCR

6 Batch Reactors Highly Degraded Manure-Media (MSL) Fed Acetic Acid Acrylic, Rod Mixed Glass, Shaker-Mixed

Acrylic, Rod Mixed

Glass, Rod-Mixed

How are Organisms Identified and Quantified in mixed culture? Each taxonomic group has ribosomal DNA (rDNA) that is unique to its group This information is available in public databases. ex: NCBI rDNA is extracted from samples quantitative Polymerase Chain Reaction assays quantify the DNA Technique requires prior knowledge of organism’s genome Probes and primers complement and bind to target DNA 10

Methods – Organism Quantification 11 1.Collect Sample 2.Extract and Purify DNA 3.Quantify the DNA of important anaerobes

Releasing DNA The key to good analysis is the total release of DNA from the cell. Use glass beads to beat cells in order to break the cell wall and membrane. DNA Needs to be released by beating with glass beads 12 Olson, UCI, CEE 263, 2008

Flowcell – Basic Principles First Cell Measures Total Gas, Second Cell Measures Methane Glass scrubber with 2 M NaOH to remove CO 2., Bromocresol Green Indicator, color transition, clear to green at pH 12

R1R2 - 5 V + Two Resistors in Series R1: 12k Ω, R2:Cd_S(1-50)k Ω 15 mC LED & 200 Ω R Gas inGas Out FLOW- CELL - + Volts

Flowcell – Wiring

2 Cell Circuit Diagram, Plan View

Biogas Chromatogram, GC

Scrubbed Biogas, no CO 2

Plumping and Circuit Diagram, 6 Reactors Physical connections to Match Software Organization R1R3R4R2R5R USB Device PC