The following four tasks correspond with each research objective as noted in the above research plan. A brief description of these tasks follows.

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1 Date of Report: June 30, 2004 EPA Agreement Number: R Center Name and Institution of Center Director: Rocky Mountain regional Hazardous Substance Research Center, Colorado State University Identifier used by Center for Project: Project No. 11 Title of Project: Assessment of Electrokinetic Injection of Amendments for Remediation of Acid Mine Drainage Investigator(s) and Institution(s): Dr. Kenneth Carlson and Dr. Amy Pruden, Colorado State University, Fort Collins, CO Type of Research: Applied Project Period: 11/30/ /01/2005 (2 years) Supplemental Keywords: electrokinetic remediation, sulfate reducing bacteria, acid mine drainage Goal of Research The goal of this research is to evaluate the technical feasibility and cost effectiveness of using electrokinetic injection delivery techniques for the remediation of acid mind drainage. Statement of Work (including objectives and tasks) The research project has four objectives: Determine the efficacy of unamended electrokinetic injection for removal of metal contaminants as oxides and hydroxides, Determine the efficacy of electrokinetic injection for delivery of organic electron donors for stabilizing metals in acid mind drainage (AMD) contamination, Characterize the microbial community that is stimulated during various electrokinetic injection process configurations and determine the metal removal efficiency of an electrokinetic injection system, and Compare order of magnitude costs and operational issues of electrokinetic injection of electron donors for AMD remediation. The following four tasks correspond with each research objective as noted in the above research plan. A brief description of these tasks follows. Task I: Determine the Efficacy of Unamended Electrokinetic Injection for the Removal of Metal Contaminants as Oxides and Hydroxides (Project Year 1) The removal of dissolved metals from AMD will be analyzed using an electrokinetic soil processing (ESP) column with only water amendments in each of the electrode wells and under different operating conditions such as current, flow rate, and metal contaminant. Task II: Determine the Efficacy of Electrokinetic for Delivery of Organic Electron Donors for Stabilizing Metals in Acid Mine Drainage (AMD) Contamination (Project Years 1 and 2)

2 The removal of dissolved metals from AMD will be analyzed using an electrokinetic soil processing column with an organic electron donor in the electrode wells and under varying operating conditions including current, flow rate, soil hydraulic conductivity, and amendment type and concentration. Task III: Characterize the Microbial Community That is Stimulated During Various Electrokinetic Injection Process Configurations and Determine the Metal Removal Efficiency of an Electrokinetic Injection System (Project Years 1 and 2) The microbial community response to the various configurations studied in Tasks II and I will be studied. Denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR) products of the community 16S ribosomal DNA will be used to characterize the microbial communities. Real time PCR will be used to determine the quantitative composition of the communities. The experiments for Task III will be similar to those described in Task II except that the soil environment will be allowed to develop biological activity. Task IV: Compare Order of Magnitude Costs and Operational Issues of Electrokinetic Injection of Electron Donors for AMD Remediation (Project Year 2) Develop a conceptual design based on experimental data and literature and estimate the capital investment and operation and maintenance costs involved in a field scale electrokinetic injection system. Costs will be compared to the costs involved with the passive permeable reactive barrier approach. Additionally, technical advantages and disadvantages of EI will be presented. Relevance of Research Electrokinetic injection for uniform delivery of amendments to the subsurface is a technology generating a significant amount of interest in the remediation of chlorinated solvents and other redox-active compounds. The deliverables from this research will include the assessment of electrokinetic delivery of amendments for acid mine drainage remediation of particular relevance to the Rocky Mountain region. Progress (for each objective/task, provide the following: (1) Progress to date; (2) % completed; (3) Problems encountered and/or unexpected results; and (4) Future activities) Task I The materials necessary for experiments in Task I have been procured. These materials include the DC power supply, electrolyte recirculation pumps, electrodes, soil, and smaller associated equipment. Additionally, the acrylic soil column and the platinum wire mesh anodes have been constructed. The DC power supply equipment and the peristaltic pumps have been calibrated and tested for functionality. Tests have been conducted on the research-quality sand and clay to determine soil classification and other basic soil properties. 2

3 The following photo depicts the first experimental electrolytical cell set-up. Two peristaltic pumps located on the upper shelf will recirculate electrolyte solution between the each solution reservoir and the electrode departments. A DC power supply provides power to the positive anode (platinum wire mesh) and to the negative cathode (titanium sheet). The soil column has 12 sampling ports on the front side of the column and six reference carbon electrodes on top of the cell for voltage drop measurements. Tracer studies are being completed to assure optimal flow through the column with out any short circuiting. (2) % completed: 20 % (3) Problems encountered/unexpected results: The procurement process for the experimental apparatus did not begin until February Long lead times on several of the items were not anticipated. As a result, the project has been active for about six months. However, all large equipment materials have been purchased and the PhD student responsible for running experiments in Task I is working full time on the research plan. (4) Future activities: Upon completion of tracer studies on the soil column, experiments using typical acid mine drainage will commence. We hope to complete this task by September Task II The soil columns and associated experimental equipment used in Task I will be used in Task II, with the addition of organic donors in the electrode wells. All large materials necessary for Task II have been purchased. (2) % completed: 0% (3) Problems encountered/unexpected results: NA. 3

4 (4) Future activities: The use of organic donors in the electrode wells will be tested once Task I has been completed. Task III All reagents have been ordered and prepared for molecular analysis of the microbial communities. The procedure for DGGE has been developed and is currently being used to assess inoculum sources for the biological column experiments. We evaluated several different inocula (from manure, wetlands and from successfully operating permeable reactive barriers) in order to determine which one will be most appropriate for starting the systems and maintaining them at high efficiency. Real time PCR is being optimized for different groups of sulfate-reducers: Desulfotomaculum, Desulfobulbus, Desulfobacterium, Desulfococcus and Desulfovibrio so that we will be able to quantify these groups of sulfate-reducers in the ESP columns. The real time PCR instrument is being calibrated to quantify these groups. Of particular note, because the Co-PI on this project (11) is also Co-PI on Project 7, we are taking the opportunity to synthesize the results from both projects. One key point of comparison is that the sulfate-reducing permeable reactive zones (SR-PRZs) being studied in project 7 rely on a highly complex microbial community, whereas we expect a much simpler community will develop in the electrokinetic systems studied in this project. It is our hypothesis that these simpler systems will be more easily controlled and less susceptible to upset. In this time prior to the setup of the biologically active experiments, we have taken the opportunity to fully characterize the microbial communities present in the complex systems so that this data will be available for comparison when the electrokinetic experiments are complete. (2) % completed: 20% (3) Problems encountered/unexpected results: (none) (4) Future activities: Once Task II is completed new ESP columns will be set up with nonsterile soil so that biological activity will develop. Two organic amendments and three different types of soil will be tested used in this task. We will then fully characterize the microbial populations which develop in these systems. Task IV This task has not yet been started. Summary of Relevant Data (if any) NA 4

5 Schedule and Scope (given the work to date, is original timetable and work plan still valid?) The original scope of work is valid, however, we may need to accelerate certain aspects of the project to achieve the original schedule. Appendix (attach a separate appendix listing publications, presentations and relevant web sites) NA 5