Accidental and intentional release of hazardous wastes threatens environmental sustainability and human health. The capacity of soils to detoxify waste has been well documented. This capacity is limited however, and natural detoxification processes often require years to restore impacted sites.
Sorption and sequestration of xenobiotic compounds within the soil matrix are critical processes affecting contaminant mobility, toxicity, and persistence. Slow desorption and release from the soil matrix to the aqueous phase represents a long-term contaminant source and hinders remediation efforts. Both synthetic and biological surfactants have been shown to enhance the apparent aqueous solubility of nonpolar organic compounds (NOC) resulting in increased bioavailability and biodegradation. However, there are also reports which suggest that some synthetic surfactants inhibit biodegradation. This inhibition is generally attributed to toxicity or reductions in bioavailability due to partitioning of contaminant into surfactant micelles. Despite conflicting data, it has been suggested that surfactants may be used to facilitate pump-and-treat and/or bioremediation of contaminated soils and aquifer sediments. Most research has focused on synthetic surfactants, but these compounds are often toxic and recalcitrant and pose the threat of additional contamination. On the other hand, biosurfactants possess similar properties to those of synthetic surfactants and have been demonstrated to enhance hydrocarbon uptake by bacteria. Biosurfactants also have the advantage of being biodegradable and can be produced on site or potentially in situ where the target contamination exists. It is reasonable to expect that biosurfactants will have similar effects on sorbed NOC, however, this aspect of biosurfactant chemistry has not been previously investigated.
The application of biosurfactants to enhance biological soil remediation requires precise knowledge of soil microbial ecology, as well as the fate and transport of NOC and biosurfactants in environmental systems. The purpose of this project is to investigate the effects of biosurfactants on the bioavailability of NOC in porous media using batch and column methodologies with real soils. Biodegradation will be evaluated with well characterized NOC-degrading enrichment cultures or native microbial populations from nonsterile soils. Biosurfactants will be obtained from well characterized glycolipid-producing pure bacterial cultures. The information acquired will be used to develop more effective remediation strategies for the reclamation of contaminated soil, vadose zone, and aquifer sediments.
The results of the proposed research will further our understanding concerning the bioavailability of hydrophobic organic compounds in soil and sedimentary environments. This knowledge will improve our ability to predict the fate of these compounds and lead to the development of more effective remediation strategies for the reclamation of contaminated soils. The interdisciplinary nature of the research requires expertise in transport phenomena, surface chemistry, microbiology, organic chemistry, and environmental engineering. The co-principal investigators have experience, expertise and interdisciplinary backgrounds that overlap to cover all of these areas. This integrated approach will insure the successful completion of the project goals
1. The Effect of Biosurfactants on the Fate and Transport of Nonpolar Organic Contaminants in Porous Media
2. Focus Catagories: GW, TRT, TS, ST
3. Key Words: Bacteria, Biodegradation, Contaminant transport, Groundwater quality, Pollution Control, Soil, Toxic substances
4. Duration: July 1, 1997 - June 30, 1999
5. Federal Funds: FY 1997 - ‘99: $69,543
6. Non-Federal Funds: FY 1997 - ‘99: $220,759
7. Mark Radosevich and Yan Jin, Department of Plant and Soil Sciences, Daniel K. Cha, Department of Civil and Environmental Engineering, University of Delaware
8. Congressional District: At Large
9. Statement of the Critical Regional or State Water Problem(s)
Accidental and intentional release of hazardous wastes threatens environmental sustainability and human health. The Northeast Region has many industrial centers where accidental or intentional releases of hazardous substances to soils and subsurface environments are common. As a result the region has numerous sites that require cleanup of soils and aquifers under various federal and state programs. The contaminated sites include government and industrial facilities (Superfund Program, approximately 50,000 across the U.S.) to leaking underground storage tanks (estimated to be several hundred thousand) . Many of the contaminated sites in this region are located in areas that have shallow water tables and course-textured, permeable soils making the groundwater more susceptible to contamination. Although the capacity of soils to detoxify waste has been well documented, this capacity is limited however, and natural detoxification processes often require years to restore impacted sites. In the United States alone, it has been estimated that hazardous waste site resoration costs may approach 1.7 trillion dollars over the next 30 years. These estimates have raised serious concerns regarding the ability to pay for site restoration. Yet in the U.S, 40 million people live within four miles of a superfund site. Therefore, it is likely that support will continue to grow for site clean-up and restoration. Consequently it is imperative that less expensive and more efficient remediation approaches be developed.
Sorption and sequestration of xenobiotic compounds within the soil matrix are critical processes affecting contaminant mobility, toxicity, and persistence. Slow desorption and release from the soil matrix to the aqueous phase represents a long-term contaminant source and hinders remediation efforts. It has been suggested that surfactants may be used to facilitate pump-and-treat and/or bioremediation of contaminated soils and aquifer sediments. Most research has focused on using synthetic surfactants to mobilize contamination for pump-and-treat systems and has shown promise in enhancing biodegradation. Biosurfactants have comparable solubilization properties of synthetic surfactants but have several additional advantages which make them superior candidates in bioremedation schemes. First, biosurfactants are biodegradable and pose no additional pollution threat. Furthermore, most studies indicate that they are non-toxic to microorganisms and therefore are unlikely to inhibit biodegradation of NOC. Biosurfactant production is less expensive, can be easily achieved ex situ at the contaminated site, and has the potential of occurring in situ. Petroleum based surfactants, on the other hand, can be toxicants, recalcitrant, and can only be derived from complex synthetic reactions making their production expensive, impossible to achieve on site, and results in the production of toxic waste byproducts. It is reasonable to expect that biosurfactants will have similar effects on sorbed nonpolar organic contaminants (NOC), however, this aspect of biosurfactant chemistry has not been previously investigated.
The results of the proposed research will further our understanding concerning the bioavailability of hydrophobic organic compounds in soil and sedimentary environments. This knowledge will improve our ability to predict the fate of these compounds and lead to the development of more effective remediation strategies for the reclamation of contaminated soils.
10. Statement of results or benefits
Reduced contaminant bioavailability due to sorption and/or diffusion through the soil matrix has already been demonstrated. These processes hinder both biologically and physically based remediation systems. Some studies have shown that both synthetic and biologically derived surfactants can enhance dispersion and bacterial uptake of hydrocarbons in surfactant-water systems while many other studies have shown an inhibitory effect. The vast majority of studies involving soil-surfactant-water systems conducted with synthetic surfactants have also shown mixed results. However, very few studies have addressed the effects of biosurfactants on the bioavailability of soil-sorbed substrates. Biosurfactants may influence these systems in several ways. First, soil solution biosurfactant concentrations above the critical micelle concentration (CMC) may enhance the overall rate of NOC degradation by: 1) enhancing the apparent solubility of NOC resulting in higher aqueous phase concentrations and thus higher rates of degradation, 2) altering the distribution of the contaminant between sorbed and solution phases, or 3) enhancing the mass transfer rate of the contaminant from the sorbed to the solution phase. Alternatively, if the micelle-associated contaminant is inaccessible to microorganisms, if the biosurfactant is toxic, or if the biosurfactant is preferentially degraded, then reduced NOC biodegradation maybe observed. Preliminary experiments have shown that treholose micelle-water partition coefficients for toluene, xylene, and trimethyl benzene were higher than those observed for either SDS or soil organic matter. Therefore, it is anticipated that the presence of biosurfactant will enhance the overall rate of NOC biodegradation via enhanced desorption. Once this has been demonstrated at the laboratory scale, the results of this research will provide the basis for developing economically and technically feasible remediation techniques based on flushing the contaminated area with biosurfactant or stimulating biosurfactant production in situ. The proposed experiments are comprehensive and will provide sufficient information to elucidate the mechanisms responsible for surfactant-enhanced NOC biodegradation, ultimately leading to the development of improved bioremediation strategies. The interdisciplinary nature of the research requires expertise in transport phenomena, surface chemistry, microbiology, organic chemistry, and environmental engineering. The co-principal investigators have experience, expertise and interdisciplinary backgrounds that overlap to cover all of these areas. This integrated approach will insure the successful completion of the project goals.
11. Nature, scope and objectives of research
Soil and aquifer remediation efforts are often hindered by contaminant sorption. Slow desorption rates limit the availability of these compounds to degradative microorganisms, hence, extend their persistence in the environment. While synthetic surfactants have been shown to enhance desorption in soils and sediments, these compounds themselves are often toxic and recalcitrant. Many biosurfactants can cause considerable reductions in surface and interfacial tensions and facilitate hydrocarbon uptake in bacteria. Biosurfactant-producing strains have also been used to enhance oil recovery. It is reasonable to expect biosurfactants to have similar effects on sorbed NOC, however, this aspect of biosurfactant chemistry has not previously been investigated.
From an ecological point of view, sorbed solutes in low nutrient environments such as soils and aquifers represent a potentially unutilized nutrient resource. An organism with the ability to locally enhance desorption of these resources could have a distinct advantage over competitors. Bacteria which are capable of degrading hydrocarbons have been shown to release surfactants for the sole purpose of facilitating the uptake of insoluble substrates. It seems plausible that organisms in low nutrient environments may use this mechanism as a survival strategy. The overall goal of this research is to assess the effect of glycolipid biosurfactants on the bioavailability of sorbed and/or sequestered NOC to soil microorganisms.
Objectives
12. Methods, procedures and facilities
Soils
Soils (Evesboro Loamy Sand and Matapeake Silt Loam) will be collected from agricultural sites in New Castle and Sussex Counties of Delaware. All soil samples will be passed through a 2 mm sieve. Their respective field moisture content will be conserved to minimize physical and chemical changes in the solid-phase organic matter. The 2 mm size fraction will be used in all sorption and biodegradation experiments. Particle-size distribution, total organic carbon content, pH, surface area, and cation exchange capacity of all sorbent materials will be determined by standard methods by The Soils Testing Laboratory of the University of Delaware.
Chemicals
Polycyclic aromatic hydrocarbons (PAH) are pollutants of major environmental concern due to their low volatility and low water solubility. PAHs are persistent and their biological transformation is believed to be a major removal process from soil systems. Phenanthrene (C14H10) will be used as a model PAH compound. It has a low water solubility of 1.3 mg/L (23) and its log Kow is 4.54 and its log Koc is 4.46. Unlabelled phenanthrene and [9-14C]-phenanthrene will be purchased from Aldrich Chemical Co. (Milwaukee, WI) and Sigma Chemical Co. (St. Louis, MO), respectively.
Bacterial Strains and Enrichment Cultures
Two different types of biosurfactants will be tested in the study: rhamnolipids typically produced by Pseudomonas sp. and trehalose produced by Nocardia sp. These biosurfactants were chosen because (i) they are both glycolipids, which are the most commonly isolated and characterized biosurfactants and (ii) both Pseudomonas and Nocardia sp. are common soil microorganisms. Rhamnolipid and trehalose will be harvested from the culture supernatants of Pseudomonas aeruginosa (American Type Culture Collection; ATCC 9027) and Nocardia erythropolis (ATCC 4277), respectively. The Pseudomonas isolate is obtainable from ATCC and the Nocardia isolate is currently maintained in our laboratory and has been used in some preliminary experiments.
Mixed populations of PAH degrading bacteria enriched from petroleum hydrocarbon contaminated sediment has been obtained from Illinois Institute of Technology (Chicago, IL). This culture will be used in the initial degradation experiments. If problems associated with the stability of this culture are encountered during biodegradation experiments, pure phenanthrene-degrading bacterial cultures will be obtained from the ATCC or from colleagues at other institutions possessing well characterized phenanthrene-degrading cultures.
Biosurfactant Production and Characterization (Cha)
Biosurfactants will be produced by growing biosurfactant-producing bacterial cultures in defined media. Preliminary experiments will be performed to optimize the media for biosurfactant production. As a starting point for optimizing media, cultures will be grown under N-limiting conditions since several studies have observed maximum biosurfactant production under these conditions. The surface tension of spent culture media in all experiments will be measured using a surface tensiometer. Once the CMC has been determined by the dilution method, the corresponding biosurfactant concentration will be determined by measuring total organic carbon (TOC) using an organic carbon analyzer.
To facilitate biosurfactant degradation and sorption assays (see description below), 14C-biosurfactants will be produced by growing biosurfactant producing bacteria in mineral salts media containing either 14C-glucose, 14C-rhamnose, or 14C-hexadecane at high specific activities. The radiolabelled substrates will be added to the culture media in a minimal carrier solvent during the late log phase to early stationary phase of growth since this is the period of the growth curve observed to coincide with the highest rate of biosurfactant production (23). The resulting radiolabelled biosurfactant will be extracted and purified as described below.
Extraction of Biosurfactants. Rhamnolipid will be recovered from the culture supernatants using centrifugation and precipitation technique outlined by Zhang and Miller (56). After removing cells by centrifugation at 6,800 x g for 20 min., rhamnolipid in the culture supernatant will be precipitated by acidification to pH 2 and recovered by centrifugation at 12,000 x g for 20 min. The precipitate will then be extracted with chloroform-ethanol (2:1) three times and the lipids will be concentrated with a rotary evaporator. Following solvent evaporation, the residue will be dissolved in 0.05 M biocarbonate (pH 8.6) for the measurement of biosurfactant concentration.
To extract the cell wall-bound biosurfactants from N. erythropolis, samples of the whole broth will be extracted with chloroform-methanol (1:2) by continuous stirring for 5 hours (36). Following the extraction, the lipids will be concentrated with a rotary evaporator to white residues (5).
Micellar Solubilization of Phenanthrene (Cha)
Two methods are commonly used to measure micellar solubilization of NOC. The first involves measuring surfactant associated increases in the apparent solubility of the NOC in saturated systems while the second utilizes a semiequilibrium dialysis (SED) approach to measure micelle-water partition coefficients (Km) at NOC concentrations below saturation. Since the degradation and transport experiments will be conducted at subsaturated phenanthrene concnetrations, the SED techique will be used in this study. SED cells with cellulose acetate membranes will be utilized to determine the distribution of the test compounds between the micellar and the aqueous phases. The membrane in a SED Cell retains surfactant micelles and their solubilized contents on one side while the surfactant monomers and unsolubilized hydrocarbons reach an equilibrium on both sides (39). SED cells will be purchased from Fisher Scientific. A cell consists of two plexiglass halves that are clamped together over a cellulose dialysis membrane (6,000 dalton MW cutoff). Aqueous solutions containing known amounts of biosurfactant and phenanthrene will be placed in retenate compartment of the cell and pure water will be added to the other side. Samples will be introduced and removed by syringe through threaded stainless steel sampling ports. After 24-hr equilibrium period, samples of the permeate compartment will be taken and analyzed to determine the concentrations of surfactant and phenanthrene. Our preliminary study based on headspace analysis indicated that the micellar partitioning of p-xylene reached an equilibrium level with two hours. Rouse et al. (39) reported that equilibrium concentration of naphthalene on both sides of the dialysis membrane was achieved in about 15 hr. A Varian HPLC with a UV detector (225 nm wavelength) will be used to determine the concentrations of phenanthrene.
Behavior of Biosurfactants in Porous Media (Radosevich and Jin)
Sorption of Biosurfactants. The efficiency with which a surfactant can affect the solubility and hence the sorption of NOCs in a soil-water system is directly influenced by the soil-water distribution of the surfactant itself. The sorptive behavior of the biosurfactants will be determined by the batch equilibration technique as well as miscible displacement through soil columns (protocol described below). Batch sorption experiments will be conducted using biosurfactant solutions and g-irradiated soils. Biosurfactant remaining in the solution phase after equilibration will be quantified by TOC analysis corrected for dissolved organic carbon released from soil. Losses of biosurfactant due to sorption to labware will be evaluated by including a set of soil-free controls. Transformation of the biosurfactants during equilibration will be minimized by using sterile soil and incubating the samples in the dark.
Possible abiotic transformation of the biosurfactants due to hydrolysis during equilibration will be determined in a preliminary experiment using radiolabelled biosurfactant. Aqueous biosurfactant concentrations and potential hydrolysis products will be monitored by radiochromatography (Beta-One, Packard Instruments). Quantification of biosurfactant concentrations will be determined using external biosurfactant standards prepared from purified culture supernatants.
Assessment of Surfactant Biodegradation. The biodegradability of the biosurfactants will be evaluated in soil microcosms by amending nonsterile soils with radiolabelled biosurfactants (as prepared above) and monitoring 14CO2 evolution over time. In the event that 14C-labelled biosurfactant production is unsuccessful, biodegradation will be evaluated in soil-water slurries by measuring aqueous biosurfactant concentrations using TOC analysis.
Mobility of Biosurfactants in Soils under Saturated Conditions. Mobility of the biosurfactants in soils will be evaluated by conducting flow-through experiments using a column system. The column will be constructed with glass (column) and stainless steel (end plates). The same soils used in batch sorption and degradation studies will be used to pack the columns. The columns will be prepared in a standardized manner and the input solution will be supplied with a peristaltic pump. Each column will be leached with deionized water to establish a steady-state flow condition prior to the experiment. A bromide (Br-) tracer experiment will be conducted for each column to test its performance and to obtain transport parameters. After the tracer experiment, the column will be flushed with 10 mM CaCl2 solution to normalize the pH and ionic strength of the system and also to be consistent with conditions used in the batch experiments. Biosurfactant solution at various concentrations (both below and above CMC) will then be introduced into the system and samples will be collected using a fraction collector at desired time intervals.
The concentrations of the Br- tracer in the outflow samples will be measured using an Accumet-25 ISE Meter equipped with a Br- electrode. The biosurfactant in the outflow samples will be quantified by analyzing TOC using an organic carbon analyzer. The breakthrough curves of the biosurfactants will be compared with the corresponding Br- breakthrough curves to evaluate biosurfactant retardation. The flow-through experimental results will be compared with results from batch systems to determine whether a traditional batch method is appropriate for supplying parameters to predict biosurfactant mobility in soils.
Biosurfactant Effects on the Fate and Transport of Phenanthrene in Soil (Radosevich/Jin)
Determination of Sorption Coefficients for Pheneanthrene in the Presence and Absence of Biosurfactants. Sorption parameters for phenanthrene in the presence of varying biosurfactant concentrations will be determined by standard batch equilibration methods. Gamma-irradiated (to prevent biological degradation during incubation), sieved soils will be reacted with phenanthrene solutions of varying concentrations. The time required for sorption equilibrium and the appropriate soil-to-solution ratio will be determined in preliminary studies. Soil-free controls will be included to monitor loss of phenanthrene through sorption to labware and other abiotic losses such as volatilization, hydrolysis, and photodecomposition.
Effect of Biosurfactants on Phenanthrene Desorption Kinetics. Desorption of phenanthrene from pre-equilibrated soils reacted for various times (in the range of 24 hours to several months) will be assessed using a batch desorption method. After equilibration, the solid and solution phases will be separated by centrifugation and the solution will be removed and replaced with an equivalent volume of 10 mM CaCl2. Sample tubes will be sacrificed periodically and desorbed phenanthrene will be measured by HPLC or liquid scintillation spectrophotometry after centrifugation. Mass balance will be completed at the conclusion of each experiment by extracting the soil with appropriate solvents to quantify the phenanthrene remaining in the sorbed state. Unextractable phenanthrene will be determined by dry combustion and capture of 14CO2. The effects of biosurfactant on desorption kinetics will be determined by performing a parallel series of experiments in which the desorption solution will contain varying concentrations of biosurfactant above and below the CMC.
Assessment of Phenanthrene Biodegradation in the Presence and Absence of Biosurfactants (Cha and Radosevich)
Degradation kinetics in Surfactant-Water Systems. The degradation kinetics of phenanthrene in solution cultures as a function of solute concentration will be determined by measuring residual phenanthrene by HPLC, and by measuring 14CO2 evolution from 14C-phenanthrene as previously described (34). The effect of varying concentrations of biosurfactant on growth and degradation will be determined by preparing culture media containing varying concentrations of the two types of biosurfactants described in the previous section. After equilibration of the phenanthrene-biosurfactant solutions, the experiment will be initiated by inoculation with a standard inoculum prepared as described in the previous section. The effect of biosurfactant on growth will be evaluated by fitting growth curves measured in the presence and absence of biosurfactant to the Monod Equation.
To evaluate the physiological effects (toxicity) of each biosurfactant on the phenanthrene-degrading cultures, growth experiments will be performed in phenanthrene-free basal salts medium containing NH4+ as an N-source and a [U-14C]-glucose as a carbon and energy source. Glucose is selected as a substrate because it is not expected to complex, sorb, or partition into biosurfactant micelles and therefore, any inhibition or enhancement of glucose mineralization in the presence of biosurfactant can be attributed to a direct effect on the physiology of the microorganisms rather than a mass transfer constraint due to the association of the substrate with the biosurfactant. The effect of biosurfactant will be evaluated by measuring rates of 14CO2 evolution.
Assessment of Phenanthrene Bioavailability in Soil-Biosurfactant-Water Systems. The effect of biosurfactants on the bioavailability of sorbed phenanthrene will be evaluated in a series of parallel degradation experiments. The soil samples will be inoculated with the microbial enrichment cultures selected for their ability to mineralize phenanthrene or an aliquot of nonsterile soil in batch serum bottle biometers. Biodegradation will be assessed by measuring 14CO2 evolution from 14C-phenanthrene. These experiments will determine the effect of desorption on concurrent phenanthrene biodegradation under otherwise standardized conditions. All treatments will be prepared in triplicate and sterilized uninoculated soils will be included as abiotic controls. The biometers will be incubated at 25±0.5oC. Cell density as a function of time will also be monitored using a most-probable-number technique or a standard plate count procedure. The mass balance of phenanthrene will be determined at the conclusion of the biometer experiments by dry combustion of the soil samples.
Effect of Biosurfactants on Retention and Degradation of Phenanthrene Under Saturated Flow Conditions (Jin)
Phenanthrene is strongly sorbed by soils (log Koc = 4.46). The possible effects of biosurfactants on phenanthrene degradation include enhancing the rates of desorption and dissolution from the sorbed phase. Therefore, the overall effectiveness of the biosurfactants can only be studied by evaluating the different rate-limiting processes, e.g., desorption, dissolution, and degradation simultaneously. We propose to conduct two sets of flow-through experiments to first evaluate the effect of the selected biosurfactants on the removal of phenanthrene sorbed to soil surfaces due to enhancement of desorption and then to study the coupled desorption and biodegradation processes simultaneously. The detailed experimental procedures are outlined in the sections that follow.
Assessment of Biosurfactant Enhanced Phenanthrene Removal. Experiments will be conducted to study the possibility of enhanced removal of phenanthrene sorbed to soil surfaces. The same flow-through system used in the biosurfactant mobility experiments will be used. The columns will be packed with sterilized soils and conditioned in the standardized manner as described previously. Each experiment will proceed in the following order:
Effect of Biosurfactants on the Coupled Desorption-Degradation Processes. Column experiements will be conducted to evaluate the overall effectiveness of the biosurfactants in their ability to enhance desorption, hence degradation of the test compounds in contaminated soils. As a common practice, saturated column experiments are conducted using deaerated input solution in order to achieve the ideal saturated flow conditions. However, such treatment causes the soil column to be anaerobic which will influence biodegradation of the test compunds. It is inappropriate to use degradation coefficients measured in batch experiments which are normally conducted under aerobic conditions to simulate results from column experiments where anaerobic conditions prevail particularly for compounds such as phenanthrene which are known to degrade only under aerobic conditions. Therefore, we propose to conduct our experiments in two distinct phases. The initial phase will involve contaminating either sterile or preinoculated soil under conditions in which degradation is minimal. The second phase will be conducted under conditions favorable for degradation. For experiments involving phenanthrene, the soil columns will be preequilibrated with degassed (anaearobic) input solution containing phenanthrene to reach a constant concentration. It is anticipated that phenanthrene-degradation will be minimal during this period. Once the column has been loaded with phenanthrene, the input solution will be replaced with the appropriate aerated input solution (see specific description of treatments below) to allow degradation to occur. Preliminary tracer experiments will be conducted with and without degassing to evaluate any possible changes in the flow conditions. A brief description of the different treatments and experimental procedure for previously contaminated soil columns is described below.
The breakthrough curve from the first column will be used to evaluate the retardation factor (R) which provides the necessary information for evaluating degradation of the test compounds in the other treatments. The results from the second and the third columns will be compared to assess the effect of biosurfactant on phenanthrene degradation.
Data Analysis. The experimental system has been constructed specifically so that the convection-dispersion equation (CDE) may be used for evaluating the outflow data that passes through the soil columns. Sorption and degradation coefficients of the test compounds will be calculated by fitting the CDE model to the measured breakthrough curves using parameters obtained from the Br-tracer experiments. This dual tracer method allows us to analyze the two parameters influenced strongly by the test compound separately from the parameters describing the water flow field, which are evaluated from the Br- signal.
Facilities
The Department of Plant and Soil Sciences at the University of Delaware is a modern research facility with a variety of instrumentation and equipment required to complete the proposed research. Facilities include: 1) Complete chromatographic capabilities (GC, GC-MS, HPLC, IC), 2) variety of spectrophotometric instrumentation (XRD, FTIR, UV-VIS, ICP, AAS, liquid scintillation counter, and 4) complete microbiological, soil physics, and molecular biology facilities. The Department of Civil and Environmental Engineering has fully equipped chemical laboratories available for use in this project. Shakers, pumps, centrifuges, automated fermentors, HPLC, liquid scintillation counter, and other routine analytical equipment are available. The department also operates an excellent machine shop for column fabrication. The only instrument currently unavailable for the successful completion of the proposed research is a dissolved organic carbon analyzer. Dr. Cha will provide matching funds ($25,000) for the purchase of a suitable instrument.
13. Related research
Removal of contaminants from the bulk aqueous phase has been shown to limit their bioavailability to microorganisms (44) and to plants (25, 31). Reduction of aqueous phase contaminant concentrations can result from a variety of processes that are primarily a function of the solubility of the contaminant and the physical and chemical properties of the soil. These processes include slow dissolution of the contaminant solid or liquid phase, sequestration in soil micropores, soil sorption (either adsorption or partitioning), and partitioning of the contaminant in a nonaqueous phase liquid.
Sorption of NOCs. Sorption is described as an equilibrium distribution of the chemical between the aqueous phase and soil organic matter. The mechanism of uptake is thought to be a partitioning phenomenon, similar to the partitioning of hydrophobic organic compound between an organic solvent phase and aqueous phase in a biphasic solvent system (7). This process is characterized by linear sorption isotherms where the sorbed concentration is directly proportional to the solution phase concentration at equilibrium. Partition coefficients are usually normalized to the mass fraction of organic carbon (foc) or organic matter (fom) of the soil to yield the organic carbon partition coefficient (Koc).
There is considerable evidence which indicates that sorption (adsorption or partitioning) can inhibit biodegradation in soils and sediments (44). Traditionally, most transport equations and fate models assume first-order biodegradation kinetics based on solution degradation rates. These models do not adequately describe biodegradation in soils or sediments and make no provisions for the rate of sorption or desorption (1). In general, description of soil biodegradation data has been improved by assuming that the compound can reside in either of two compartments one which is considered to be accessible to microorganisms, and one which is inaccessible (15). After the substrate in the accessible compartment is degraded, the rate of biodegradation is determined by the rate of desorption or diffusion of the substrate in the inaccessible pool to the accessible pool. This approach assumes that the rate of transfer between the two compartments and the rate of conversion of the substrate to products are controlled by first-order kinetics. Ogram et al. (30) successfully modeled 2,4-D biodegradation by assuming that sorbed 2,4-D was completely protected from biological degradation and that sorbed and solution phase bacteria degraded solution phase 2,4-D with equal efficiency.
Surface adsorption can also limit biodegradation. Miller and Alexander (27) observed slower rates of benzylamine degradation by a pure bacterial culture in the presence of montmorillonite than in the absence of clay. Smith et al. (45) used fluorescence spectroscopy to independently monitor changes in solution and surface-bound quinoline concentration during biodegradation in a smectite clay suspension. Utilization of solution quinoline was 30 times faster than bound quinoline and overall microbial utilization of quinoline was desorption-rate limiting.
Despite the mixed success in modeling sorption limited biodegradation in soils and sediments the available data indicates there may be some validity to the concept of available and unavailable compartments. If so, substrates localized in the unavailable compartment(s) may be an unexploited resource. But it is equally reasonable to expect the existence of microorganisms that are equipped in some way to exploit this resource. Evidence supporting this concept has been obtained recently (12) in a study of the bioavailability of sorbed naphthalene to two distinctly different bacterial species capable of degrading naphthalene. Bioavailability assays gave dramatically different results for the two bacterial species. For one unidentified soil-isolate, designated NP-ALK, desorption clearly limited both the extent and rate of naphthalene degradation. However, for the other species, Pseudomonas putida ATCC 17484, both the rate and extent of degradation exceeded predicted values and resulted in enhanced rates of naphthalene desorption from soils. P. putida 17484 was noted to be chemotactic towards naphthalene, and adsorbed to surfaces reversibly. It appears likely that these attributes may be involved in the ability of this organism to utilize sorbed naphthalene. It was proposed by the authors that strain 17484 was able to mineralize surface localized, labile sorbed naphthalene establishing a steep concentration gradient to promote desorption of non labile sorbed naphthalene. NP-ALK, on the other hand relied strictly on passive desorption of bound naphthalene. The results of study suggests that limitations to biodegradation due to sorption may be species dependent, and suggests that cellular adsorption may allow some species to utilize sorbed solutes.
Slow Diffusion Through the Soil Matrix. The uptake of NOC by soil is usually characterized by a two-stage approach to equilibrium: a short initial stage of rapid uptake lasting for minutes to hours followed by an extended slower stage lasting days or months (4). The slower rate-limiting stage is generally attributed to diffusion of the solute molecules through soil structures such as water films, intra-aggregate micropores, and the three-dimensional matrix of soil organic matter. Diffusion of solute molecules into increasingly more remote sites within the soil matrix can continue over time (32, 33). The overriding feature of this process is that with aging the fraction of solute molecules in the slowly reversible sorbed state (resistant fraction) increases relative to the fraction in the bulk aqueous phase or the more rapidly reversible (labile) fraction. Therefore, it is reasonable to expect residence time (i.e., the time a contaminant is in contact with soil) to have an influence on the rate of desorption and subsequently on biodegradation. Steinberg et al. (46) showed that aged residues of 1,2-dibromoethane in field soil samples were much less biodegradable than freshly added 14C-labeled compound in the same samples. Residues of 1,2-dibromoethane persisted for years despite being highly volatile and biodegradable by soil microorganisms. Similar results were observed for simazine taken from agricultural soil with a long term history of simazine treatment (42). In more controlled laboratory studies, bioavailability in soils inoculated with microbial cultures capable of degrading test chemicals, has been shown to decline with chemical aging for naphthalene (13), phenanthrene and p-nitrophenol (17), and atrazine (35). In some studies, physical disruption of the soil structure enhances chemical availability suggesting that a fraction of the chemical was protected from microbial attack through sequestration in soil micropores (17, 46).
Mihelcic and Luthy (26) used a retarded intra-aggregate radial diffusion model (RDM) that described sorptive constraints on the biodegradation rate of naphthalene. Similar results were obtained for the biodegradation of a-hexachlorocyclohexane (37). These authors applied both a first-order model and the RDM to the desorption and bioconversion data. In the case of a-hexachlorocyclohexane, the RDM did not provide significant improvement in describing the data over the simpler first-order model. In fact, the biodegradation data could only be described by the RDM if penetration of the microorganisms into the interior of the aggregates was assumed. The effect of retarded intra-aggregate diffusion (partitioning plus diffusion through micropores becomes more pronounced for more hydrophobic compounds since the sorption/desorption kinetics are expected to be slower (54). Scow and Alexander (41), using a diffusion based model, showed that clay aggregates reduced the concentration of available p-nitrophenol and lowered the apparent biodegradation rate constant.
Influence of Surfactants on Bioavailability
Micellar Solubilization. "Surfactants" or surface active agents are compounds with both hydrophilic and hydrophobic moieties with the ability to reduce surface and interfacial tensions and enhance the aqueous solubility’s of relatively water insoluble compounds. Synthetic surfactants have been used by the oil industry to enhance oil recovery for many years (3, 43), and also by pesticide manufacturers to formulate hydrophobic pesticides. However, the application of surfactants to enhance desorption or dissolution of pollutants from contaminated soils, sediments, and aquifers has only recently received attention (51). The solubilization of NOCs by surfactant micelles can be defined by micelle-water partition coefficients (Km). Relationships between Km and other measures of hydrophobicity (octanol-water partition coefficient (Kow), water solubility, and molecular surface areas) have been investigated (20, 50). The log Km values of eleven nonionic organic solutes in sodium dodecyl sulfate (SDS) surfactant micelles were determined and found to be linearly related to log Kow, log aqueous solubility, and molecular surface area. The strongest predicting parameter of Km was found to be Kow (50). This relationship has obvious implications for predicting the mobilization of these compounds in the presence of solubilizing surfactants. Valsaraj and Thibodeaux (50) provided a modified equation for the retardation factor (R) in the presence of surfactant micelles.
Where n is the porosity of the soil, r is the bulk soil density (g cm-3), vm is the molar volume of water (l mol-1), and C is the total surfactant concentration (mol l-1). The enhanced solubilization of NOC is most dramatic for compounds with large Kow values. As a result the effect of surfactants on the retardation factor is negligible for compounds with low Kow.
The effectiveness of the anionic surfactant sodium dodecyl sulfate (SDS) in solubilizing PAH compounds from soil and sediment slurries was examined by Jafvert (19). SDS was very effective in solubilizing PAH. Calculated and experimental distribution coefficients indicated that on an organic carbon normalized basis SDS micelles had a similar affinity (sorption potential) as soil organic matter for these compounds. In one experiment in which phenanthrene was pre-incubated with the solid phase, SDS micelles were effective in recovering sorbed phenanthrene. At concentrations of surfactant between 10-30 mM essentially 100% of phenanthrene was recovered in the aqueous phase. Water solubility enhancements of DDT and trichlorobenzene (TCB) by triton series, SDS, cetyltrimethylammonium, and Brij 35 was studied at surfactant concentrations above and below the CMC (20). All surfactants increased the aqueous solubility of the compounds tested. Solubility curves increased slightly with increasing surfactant concentration below the CMC, and turned sharply upward at concentrations above the CMC (20).
Surfactant selection criteria for maximum solubility enhancement has been the subject of several studies (3, 51). One such study examined the effect of a variety of surfactants on the solubility and desorption enhancement of anthracene and biphenyl from artificial soil (51). Surface tension minimization, CMC, hydrophile-lipophile balance number (HLB), solubilization efficiency, and partition coefficient of the surfactant for soil were examined and found to be useful but any one parameter by itself was inadequate to predict surfactant efficiency. Choosing a surfactant with minimal adsorption was a critical parameter. In more recent studies, triton series surfactants were shown to enhance desorption of trichloroethylene from soils and aquifer sediments (9, 47). The available data suggest that synthetic surfactants can dramatically influence the sorption behavior of NOC in soils and aquifer sediments, but comparable data does not exist for biosurfactants.
Preliminary Results. Recent work in our laboratory has investigated the potential application of non-toxic biosurfactants produced from Nocardia erythropolis (ATCC 4277) for enhancing solubility of monoaromatic hydrocarbon compounds. Solubilization potential of biosurfactant micelles was quantified by determining the Km for toluene, p-xylene and trimethylbenzene. N. erythropolis was grown in a 500 mL batch reactor with n-hexadecane as the sole carbon source to produce biosurfactants.
The production of biosurfactants was confirmed by the lowering of surface tension in the culture broth to 41 dynes/cm in 24 hours. Surface tension reached a minimum (35.2 dynes/cm) within 4 days after inoculation. This result suggested that the biosurfactants produced by N. erythropolis exhibited comparable surface-active characteristics to synthetic surfactants which typically reduce surface tension to 30-35 dynes/cm. Surfactant concnetration was determined by gradually diluting a known volume of culture medium while monitoring the surface tension.
The Km values of toluene, p-xylene, and trimethyl benzene were obtained from the slope of isotherm plots (Fig. 1). The results showed that the degree of NOC partitioning was related to solubility of the compound. The biosurfactant had the greatest affinity for trimethylbenzene (solubility = 57 mg/L) followed by p-xylene (solubility = 185 mg/L) and toluene (515 mg/L). The partition constants were: KToluene = 0.0020 L/mg, Kp-Xylene = 0.0086 L/mg, KTMB = 0.0412 L/mg (not shown in Fig. 1). The biosurfactant partition coefficient of p-xylene determined in our study was an order of magnitude greater than that of synthetic surfactant SDS (KSDS = 0.000386 L/mg) determined by Holsen et al. (18) using the same methodology. This indicates that biosurfactants produced from N. erythropolis may be much more efficient in enhancing the solubility of NOCs than commonly utilized synthetic surfactants.
Figure 2 compares our experimental results on the relationship between the calculated partition coefficient and the log solubility of each compound to the predicted relationships between Koc and solubility presented by Hassett et al. (16) for soil organic matter. The results indicate that biosurfactants can partition greater amounts of NOC than an equivalent amount of soil organic matter.
Biodegradation in Surfactant-Soil-Water Systems. Numerous studies have investigated the role of synthetic surfactants in the biodegradation of nonaqueous phase liquids and other xenobiotic chemicals in soils. Enhancement and inhibition of biodegradation in the presence of commercial surfactants have been observed. The mechanisms responsible for these observations are poorly understood. In general, enhanced degradation is attributed to enhanced solubility (6, 48), an increase in the interfacial surface area between immiscible fluids (38), or enhanced desorption (2). Inhibition of biodegradation can result from adverse interaction of surfactants with cell membranes resulting in cellular toxicity or sequestration of the contaminant within surfactant micelles. Laha and Luthy (22) observed an inhibition in phenanthrene mineralization from soil-slurries in the presence of nonionic surfactants above the CMC. When the surfactant concentration was subsequently diluted below the CMC the inhibitory effect was reversed suggesting that cell lysis was not involved. Modeling of the phase partitioning of the phenanthrene in the soil-surfactant-water system showed that reductions in the equilibrium aqueous phase phenanthrene concentration due to sorption and micellization were insufficient to explain the observed inhibition in mineralization. Similar results were obtained by Guha and Jaffé (14) for phenanthrene degradation in the presence of Triton N101, Triton X100, and Brij 35. Modeling efforts indicated that the bioavailable fraction decreased with increasing surfactant concentration above the CMC. Complete inhibition was observed in the presence of Brij 35 micelles. The authors attributed differences in bioavailability of the micellar phase to differences in polyoxyethylene chain length suggesting that longer, more hydrophillic chains of Brij 35 may have hindered the interaction between micelles and hydrophobic bacteria preventing direct phenanthrene uptake from the micellar phase. Rouse et al. (39) noted a correlation between inhibited biodegradation of hydrocarbons and surfactant concentrations above the CMC when mixed microbial cultures were used as the catalytic agents. It is important to note that the vast majority of studies have focused on commercial surfactants and very little is known regarding the behavior of biosurfactants in soils (39).
Enhanced Degradation of N-Alkanes in the Presence of Biosurfactants. Biosurfactants are produced by microorganisms for a variety of reasons. It has been noted for many strains that surfactant production is often associated with growth on carbon-rich, hydrophobic substrates such as oil or paraffin. It is believed that secretion of the surfactant lowers the interfacial tension thereby making the substrate more readily available (8, 10).
The importance of water-soluble fraction on the degradation of nonaqueous phase liquids (NAPL) has been demonstrated in mineralization studies of octadecane (21, 49) and degradation studies of tri-, tetra-, penta-, hexa-, and octadecane (28). The linear growth (or mineralization) kinetics observed coincided with a linear rate of compound dissolution. Growth rate of Pseudomonas sp. in the presence of liquid diphenylmethane, however, was greater than the growth rate of the same bacteria observed in the presence of solid diphenylmethane (53). This difference was attributed to the microbial utilization of liquid hydrocarbon at the aqueous-hydrocarbon interface. Goswami and Singh (11) showed that hexadecane-grown Pseudomonas cells have surface properties that facilitate direct attachment of cells to hydrocarbon droplets.
The available data on biosurfactants suggests that they may be very effective in enhancing the solubility of many nonpolar organic compounds. For example, rhamnolipid biosurfactant produced by Pseudomanas aeruginosa enhanced the dispersion and subsequent biodegradation of octadecane in solution cultures (56). It is reasonable to expect biosurfactants to also enhance desorption and dissolution of sorbed NOC and NAPL, respectively and thus have potential application in enhancing remediation of contaminated soils and sediments. Little information exists, however on the ability of biosurfactants to enhance the solubility of NOC and/or enhance their desorption from sorbent materials.
Relation to Long-Term Project Goals
Elucidating the effect of rate-limited processes on the fate and transport of xenobiotic chemicals in soils has been and will continue to be a major research focus for all three principal investigators. As our understanding of the biological, chemical, and physical processes governing the environmental fate of xenobiotics improves, it will be possible to engineer more effective remediation systems for detoxifying contaminated soil and groundwater. Since these processes are intimately connected, multidisciplinary approaches are critical to the accomplishment of this goal. It is important to note that the combined expertise of this research team very effectively addresses the multidisciplinary nature of the proposed research. If the results of the proposed research indicate that biosurfactants can enhance the overall rate of NOC biodegradation then future research efforts will focus on evaluating the beneficial effects at the field scale. One limitation to this approach involves the delivery of the biosurfactant to contaminated areas, particularly in the unsaturated zone. This limitation could be overcome by stimulating in situ biosurfactant production by indigenous microbial populations. Existing research suggests that in many strains, biosurfactant production is very strictly regulated and associated with nitrogen metabolism with the highest levels of biosurfactant being produced under N-limiting conditions (21, 29). Current bioremediation strategies for hydrocarbon contaminated sites usually involve addition of nitrogen and phosphorus fertilizers to stimulate growth of hydrocarbon-degrading microorganisms, a practice that may inhibit biosurfactant production and thus subsequent hydrocarbon utilization. Significant enhancement in hydrocarbon utilization at these sites might be achieved by stimulating and/or deregulating biosurfactant producing operons in environments containing excess nitrogen levels. Future research will focus on the effective delivery and/or stimulation of in situ biosurfactant production as well as the effect of biosurfactants on the degradation of NAPL and NOC in NAPL-soil-water systems.
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Mark Radosevich
Department of Plant and Soil Sciences, 149 Townsend Hall
University of Delaware, Newark, Delaware 19717-1303
Telephone: (302) 831-1376 (w) Fax: (302) 831-3651
E-mail: mrad@.udel.edu
Education
M.S., Microbiology, Colorado State University, 1990
California Teaching Certificate, National University, 1987
B.S., Bacteriology, University of California, Davis, 1986
Professional Experience
Publications
Radosevich, M., S.J. Traina, and O.H. Tuovinen. 1996. Biodegradation of atrazine in surface soils and subsurface sediments collected from an agricultural research farm. Biodegradation. 7:137-149.
Radosevich, M., S.J. Traina, and O.H. Tuovinen. 1995 Degradation of binary and ternary mixtures of s-triazines by a soil bacterial isolate. J. Environ. Sci. and Health part B. 30:457-477.
Radosevich, M., S.J. Traina, and O.H. Tuovinen. "s-Triazine Biodegradation." U.S. Patent Number: 5,429,949 United States Patent Office. July 4, 1995.
Radosevich, M., Y.-L. Hao, S.J. Traina, and O.H. Tuovinen. 1995. Degradation and mineralization of atrazine by a soil bacterial isolate. Appl. Environ. Microbiol. 61:297-302
Radosevich, M., J.J. Crawford, S.J. Traina, K.-H. Oh, and O.H. Tuovinen. 1993. Biodegradation of atrazine and alachlor in subsurface sediments. p. 33-41. In Sorption and degradation of pesticides and organic chemicals in soil (D.M. Linn, ed.), Special Publication No. 32, Soil Science Society of America, Madison, WI.
Sims, G.K., M. Radosevich, X.T. He, and S.J. Traina. 1991. The effects of sorption on the bioavailability of pesticides. p. 119-137. In Biodegradation: natural and synthetic materials (W.B. Bettes, ed.), Springer-Verlag London Ltd., London, UK.
Radosevich, M. and D.A. Klein. 1991. Bacterial enumeration and mercury volatilization in deep subsurface sediment samples. Bull. Environ. Contam. Toxicol. 51:226-233.
Funding
Radosevich, M. 1994 - 96. The effect of soil residence time and dissolved organic carbon amendment on the sorption-limited biodegradation of atrazine. United States Department of Agriculture, CSRS/NRICGP. Funds awarded: $82,400.
Radosevich, M., Q. Johnson, and B. Vasilas. 1995-96. Effect of poultry manure on the efficacy of soybean herbicides. Delaware Soybean Board. Funds awarded: $7,500.
Johnson, Q., B. Vasilas, M. VanGessel, and M. Radosevich. 1996-97. Effect of poultry manure on the efficacy of soybean herbicides. Delaware Soybean Board. Funds Awarded: $9,080. (continuation of ‘95-’96 grant)
Radosevich, M. and J.J. Fuhrmann. 1996-97. Mechanisms of enhanced organic contaminant degradation in rhizosphere soil. College of Agricultural Sciences Competitive Grants Program. Funds Awarded: $12,130.
Radosevich M. and S. Kitto. 1995-96. Goals 2000: Development of curriculum integration modules and distance learning network. Kennett Consolidated School District and The State of Pennsylvania. Funds Awarded: $462,100 (University of Delaware sub-contract: $30,000).
DANIEL K. CHA, Co-Principal Investigator
EDUCATION
Ph.D. University of California, Berkeley - Civil Engineering - 1990
M.A.Sc. University of British Columbia - Bio-Resources Engineering - 1986
B.Sc. McGill University - Agricultural Engineering (Great Distinction) - 1984
PROFESSIONAL EXPERIENCE
93 - 95 Assistant Professor, Environmental Engineering, Illinois Institute of Technology, Chicago, IL.
91 - 93 Visiting Assistant Professor, Environmental Engineering, Illinois Institute of Technology.
90 - 91 Consulting Engineer, Novatec Consultants Inc., Vancouver, B.C.
87 - 90 Engineering Trainee, Sacramento Wastewater Treatment Plant, Sacramento, CA.
PUBLICATIONS
I. Refereed Publications
McCue, J. J., Holsen, T. M., Cha, D. K., Gauger, W. K., and Kelley, R. "Effect of amorphous ferrous sulfide on the microbial reductive dechlorination of PCB Aroclor 1242." In Press, Environmental Toxicology and Chemistry (1996).
Moschandreas, D. J., Cha, D. K. and Qian, J. ìMeasurement of indoor bioaerosol concentrations by a direct counting method.î In Press, Journal of Environmental Engineering, ASCE (1996).
Cha, D. K., Song, J. S., Sarr, D. and Kim, B. J. ìHazardous waste treatment technologies.î Accepted for publication, Water Environment Research (1996).
Su, M. C., Cha, D. K., and Anderson, P. R. "Influence of selector technology on heavy metal removal by activated sludge: secondary effects of selector technology." Water Research, 29, 971 (1995).
Cha, D. K., Jenkins, D., Lewis, W. P., Kido, W. H. "Process control factors influencing Nocardia populations in activated sludge." Water Environ. Research, 64, 37 (1991).
Park, A. J., Cha, D. K., and Holsen, T. M. ìEnhancing solubilization of sparingly soluble organic compounds by biosurfactants produced from Nocardia erythropolis.î Submitted for publication, Water Environ. Research.
II. Technical Reports
Kim, B. J., Cha, D. K., and Song, J. S. ìBiotechnology to separate and treat metals in sludge and wastewater.î USACERL Technical Report 95/44, US Army Corps of Engineers (1995).
Zenz, D. R., Cha, D. K., and Moschandreas, D. J. ìBioaerosol emissions from aeration tanks: literature review.î Submitted to New York City Department of Environmental Protection (1995).
Moschandreas, D. J. and Cha, D. K. ìAnalysis of volatile organic compound emissions from the New York city wastewater collection/treatment system.î Final Report submitted to New York City Department of Environmental Protection (1995).
RESEARCH GRANTS AND CONTRACTS
| 1996-1997 | Bioavailability of Copper: Effects of Chemical Speciationî International Copper Association, $ 163,319 (co-PI) |
| 1995-1997 | Enhancing biodegradation of sorbed hydrophobic compounds using nonionic surfactants.î Great Lakes Mid-Atlantic Hazardous Substance Research Center, $ 140,207 (co-PI). |
| 1995 | Influence of aerobic selector on the population dynamics of activated sludge process.î Hammond Sanitary District, $ 5,000 (PI). |
| 1993-1994 | Bioleaching of heavy metals from contaminated solids.î U. S. Army Construction Engineering Research Laboratories, $ 24,999 (PI). |
| 1994 | ìMicrobial stabilization of pyritic mine gob tailings.î Institute of Gas Technology, $ 4,000 (PI). |
| 1992-1993 | Evaluation of anaerobic filter for the treatment of candy company wastewater.î Ferrara Pan Candy Co, $ 21,000 (PI). |
| 1992 | Degradation of naphthalene by Nocardia amarae.î Illinois Institute of Technology, $ 10,000 (PI). |
Yan Jin
Department of Plant and Soil Sciences
University of Delaware,
Newark, DE 19717-1303
Tel:(302) 831-6962 Fax: (302) 831-3651
E-mail: yjin@pollux.udel.edu
Education
Dissertation: Transport and Transformations of Volatile Organic Chemicals in Unsaturated Soils, Advisor: Dr. William A. Jury
M.S. in Soil Chemistry, New Mexico State University, 1989
Thesis: Toluene Behavior in Sludge-Amended Soils, Advisor: Dr. George A. O'Connor
B.S. in Soil Science, Agricultural University of Hebei, P R China, 1983
Research Interests, Experiences and Skills
Publications
Current Grant Support
15. Training Potential
Funding will be used to support one Ph.D. student on an annual basis. An additional Ph.D student supported on non-federal funds will also work on the project. It is anticipated that several undergraduates will also be involved in the research through independent study courses and hourly wage programs.
Year 1 |
Year 2 |
|
||||||||||
| Budget Category | Federal | Non-Federal | Federal | Non-Federal | Federal | Non-Federal | ||||||
| Personnel | ||||||||||||
| PI's | ||||||||||||
| Mark Radosevich | $0 |
$8,521 |
$0 |
$8,862 |
$0 |
$17,383 |
||||||
| Yan Jin | $0 |
$8,850 |
$0 |
$9,204 |
$0 |
$18,054 |
||||||
| Daniel K. Cha | $0 |
$13,371 |
$0 |
$13,906 |
$0 |
$27,277 |
||||||
| Graduate Research Assistant (1) | $12,000 |
$0 |
$12,480 |
$0 |
$24,480 |
$0 |
||||||
| Graduate Research Assistant (1) | $0 |
$12,000 |
$0 |
$12,480 |
$0 |
$24,480 |
||||||
| Fringe Benefits | $0 |
$8,915 |
0 |
$9,272 |
$0 |
$18,187 |
||||||
| Equipment | ||||||||||||
| TOC Analyzer | $0 |
$25,000 |
$0 |
$0 |
$0 |
$25,000 |
||||||
| Expendable Materials and Supplies | $8,000 |
$0 |
$8,000 |
$0 |
$16,000 |
$0 |
||||||
| Travel | $3,000 |
$0 |
$3,000 |
$0 |
$6,000 |
$0 |
||||||
| Tuition | $11,250 |
$11,250 |
$11,813 |
$11,813 |
$23,063 |
$23,063 |
||||||
| Total Direct Costs | $34,250 |
$87,907 |
$35,293 |
$65,537 |
$69,543 |
$153,444 |
||||||
| Indirect cost of Federal TDC (34%) | $0 |
$3,740 |
$0 |
$3,740 |
$0 |
$7,480 |
||||||
| Indirect Costs Non-Fed. (34%) excluding equipment |
$0 |
$13,483 |
$0 |
$14,023 |
$0 |
$27,506 |
||||||
| Total Cost | $34,250 |
$105,130 |
$35,293 |
$83,300 |
$69,543 |
$188,430 |
||||||
BUDGET BREAKDOWN 1997-1998
Project Number:
Principal Investigator: Dr. Mark Radosevich
Cooperating Investigators Dr. Yan Jin and Dr. Daniel K. Cha
Year 1
| Cost Category | Federal |
Non-Federal |
Total |
| 1. Salaries and Wages | |||
|
0 |
$8,521 |
$8,521 |