Oxford University
Civil Engineering
Department of Engineering Science
Using Electrokinetics and Bioaugmentation to Optimise the Remediation of PCP contaminated Soil
Gavin Lear,
Department of Engineering Science,
University of Oxford,
Parks Road,
Oxford,
OX13PJ,
UK. Tel: +44(0)1865 273 167 Fax: +44(0)1865 273 907.
E-mail: gavin.lear@eng.ox.ac.uk
Introduction
There is increasing demand for cheaper and more effective in-situ technologies to enable the remediation of contaminated land. Electrokinetics provides a novel method for the extraction of organic chemicals from contaminated sites, stimulating a directed movement of contaminant in response to the presence of an electric current. Recent studies have sought to combine the processes of electrokinetics and bioremediation in order to improve the clean-up process. Bioaugmentation is the method of inoculating a site with degrading organisms in order to increase rates of contaminant deterioration. However, the effectiveness of bioremediation alone depends on the ability of the degrading microorganisms to gain access to the contaminants. By combining bioaugmentation and electrokinetics it is hoped to increase the contact frequency between both the contaminants and soil microbial community, optimising the remediation process. It is the aim of this study to develop a better understanding of the interactions between the soil, a pentachlorophenol contaminant, and both the indigenous and specific bacterial inocula communities following the application of electrokinetic technology.
Degradation of PCP in Liquid Media
A range of PCP degrading micro-organisms have been screened for their ability to degrade PCP, with Sphingomonas sp. UG30 determined most efficient under the conditions used. Sphingomonas sp. UG30 has the ability to fully degrade up to 300mM PCP within 3 hours when supplemented with extra carbon (Sodium-glutamate), and is able to degrade PCP in soil, with the optimal degradation rate determined using a PCP concentration of about 75 mg/kg dry soil.
Figure 1: Degradation of PCP by Sphingomonas sp. UG30 (UG30) in MS-glutamate medium with 250mM PCP. Data are (above) optical density Log10(OD600nm)+2, (below) PCP concentration ((m M) (s )U+250mM PCP;(6 ) U-250mM PCP; (l )250mM PCP and chloride ion concentration (mM) (s )U + 250mM PCP (l ) 250mM PCP.
Electrokinetics
Unlike when tested in liquid media, in many cases the full degradation of a pollutant in a soil is not achieved. Within a soil, some of the PCP is inevitably contained within pores too small for bacterial interaction, or perhaps it is bound to residues in a form that makes it unavailable to the bacteria. For example, contaminant molecules contained within nanopores with diameters <100nm are probably unavailable to soil bacteria (Nam & Alexander, 1989) as their cells are too large (³ 0.2m m) for bacterial entry. We would thus say that the pollutant is not 'bio-available', and it is with the application of electrokinetics that we aim to overcome this phenomena.
The occurrence of electrokinetics transpires when soil is electrically
charged with (usually) a direct electric current. Electrodes are introduced
to the soil and the current produces hydrogen ions at the anode and hydroxyl
ions at the cathode, which migrate into the soil, with a resulting pH
gradient. Furthermore, this low-level electric current results in both
physiological and hydrological changes in the porous soil mass, allowing
the directional transport of particles and ions. Theoretically then,
this technology could be utilised to move pollutants from locations in
a soil where they are not available to microbial degraders (i.e. to increase
bioavailability,as illustrated below).
(a) After bioaugmentation, some of the pollutant remains unavailable
for bacterial mineralisation due to its position within or adsorption
to soil particles.
(b) Electrokinetics induces the movement of a pollutant as a result of
their charge and orientation within the electric field, allowing bacterial
degradation.
Figure 2: Schematic representation of the increasing bioavailability
of an aromatic hydrocarbon before (a) and after (b) the application of
electrokinetics
The mechanisms which cause the transport of the contaminant under the action of an electric field may be subdivided as electromigration, electroosmosis and electrophoresis. explanation of these processes.
Current and Future Work
General
Work so far has focused on developing the techniques and expertise needed to combine the processes of electrokinetics and bioaugmentation. The biodegradation of PCP in soil by Sphingomonas sp. UG30 has been achieved under a range of now optimised conditions (i.e. pH, soil water content, temperature and pollutant age) and their presence and viability within the soil monitored by molecular (MPN/PCR) techniques.
Further work will aim to combine the process of bacterial inoculation with electrokinetics in order to optimise this union of technologies to allow the greatest rates of contaminant degradation. A range of situations may be varied, such as current strength, current pulsing, using different inoculation strategies etc, in order to improve the process. At all times, increases in pollutant bioavailability will be closely monitored, in order to begin to understand to what extent applying electrokinetics changes the contact frequency between the bacteria and pollutant.
Monitoring Bioavailability
A molecular tool to allow a direct measurement of bioavailability is now being developed using PCP inducible genes within the degradation pathway. In essence, we may monitor the contact rate between the microbial degraders and pollutant as the bacteria themselves create a signal (mRNA) when they come into contact with PCP, enabling changes in bioavailability to be observed directly on a very small scale. Similarly, extraction techniques are being developed which are hoped to remove similar amounts of the PCP from the soil as mineralising bacteria, thereby giving us a cheap, rapid extraction technique to monitor changes in bioavailability.
Bacterial Movement and Survival under the Influence of an Electric Current
Bacteria themselves may act as charged particles under the influence of the electric field, moving towards the cathode as a result of electroosmosis and towards the anode as a result of their negative surface charge. Currently studies of being undertaken of a range of bacteria with different characteristics to determine how bacteria react to the application of electrokinetics and what factors influence their survival and movement when exposed to this process.
Mycobacteria |
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A greater bacterial movement has so far been observed when applying an electric current to the soil as compared to a control, but the movement is not biased towards either electrode (below).
Figure 3: Normalised bacterial (Pseudomonas fluorescens SBW25) cfu's within an electrokinetic cell, as compared to a control cell with no current added. Samples are taken at a range distances from the point of inoculation, over the course of one week. (n.b The anode is to the right of the x-axis)
Isolation of New PCP Degraders
An enrichment process is currently being used as a method to determine new PCP degrading isolates present within PCP contaminated soils. In essence, the enrichment process involves continually re-inoculating a minimal salts growth media, with soil and various chemical inducers, into media containing higher amounts of PCP, gradually selecting those bacteria better able to withstand high PCP concentrations. A pH indicator method is then used as a crude way of determining which isolates may have PCP degrading capabilities, as the process of PCP de-chlorination decreases the pH of the growth media, causing a resultant colour change. Isolates which produce positive results using the indicator technique are then tested more exhaustively until their ability to degrade PCP may be fully verified.
Effect of PCP on Background Soil Microbial Populations
Little research has so far been conducted regarding the effect of the application of electrokinetics. A number of studies are intended to be conducted to determine how both the background microbial population and the bacterial inocula are affected by the application of an electric current, examining factors such as changes in bacterial community structure and following the survival of a range of bacterial functional groups. Soil microbial respiration and enzyme activity may similarly be studied, and marked bacteria inoculated within the soil be used to examine the fate of individual bacterial types.
References
McGrath, R and Singleton, I (2000) Pentachlorophenol transformation in soil: a toxicological assessment. Soil Biology and Biochemistry 32:1311-1314
Nam, K and Alexander, M (1998) Role of nanoporosity and hydrophobicity in sequestration and bioavailability: Tests with model soils. Environmental Science and Technology. 32: 71-74
Pignatello, JJ., Martinson, MM., Steirt, JG., Carlson, RE and Crawford, RL (1983) Biodegradation and photolysis of pentachlorophenol in artificial freshwater systems. Applied and Environmental Microbiology 46 (5):1024-1031
Seiler, JP (1991) Pentachlorophenol. Mutational Research 257: 24-27
U.S. EPA (1997) Treatment technology performance and cost data for remediation of wood preserving sites. EPA/625/R-97/009. Washington, D.C.
Wang, YJ., Ho, YS., Jeng, JH., Su, HJ and Lee, CC (2000) Different cell death mechanisms and gene expression in human cells induced by pentachlorophenol and its major metabolite tetrachlorohydroquinone. Chemico-Biological Interactions 128(128): 173-188
