REMEDIATION OF PETROLEUM CONTAMINATED SOILS - SECTION 5 potx

The most important soil factors that affect degradation are water; temperature; soil pH; aeration oroxygen supply; available nutrients, i.e., nitrogen N, phosphorus P, potassium K, sulfu

Section 5

Optimization of Bioremediation

There are three key issues that should be addressed to achieve a successful bioremediation (Tiedje,1993) It must be determined whether the contaminant is biodegradable, whether the environment ishabitable (presence of toxic chemicals or sufficient life-sustaining growth factors), and what the rate-limiting factor is and whether it can be modified In an ecological approach to bioremediation, theimportant issue is to establish whether or not the conditions of natural selection can be expected to bemet within the site vicinity With this approach, more emphasis is placed on meeting requirements ofthe microorganisms, and less on measuring the actual pollutant

Each contaminated site exhibits different characteristics and requires a site-specific remediation plan(Forsyth, Tsao, and Bleam, 1995) Decontaminating a site polluted with hazardous materials is a complexprocedure involving systematic, step-by-step problem solving The conditions necessary to optimize theefficiency of microbial systems in degrading environmental pollutants and the economics required must

be assessed to select and implement cost-effective biotreatment This requires understanding of themicroorganisms and the conditions necessary for them to become established and maintained, and thescientific data must be translated into cost-effective, full-scale cleanup processes Augmentation withproven contaminant-degrading microorganisms can save time and money over alternative approaches.This section discusses how optimum conditions for bioremediation can be achieved through sitemanipulation, biological intervention, or chemical treatment It is important to monitor the process todetermine that biodegradation is occurring and to allow the conditions to be modified as necessary, tomaintain optimum performance

The various chemical and physical properties of a soil determine the nature of the environment in whichmicroorganisms are found (Parr, Sikora, and Burge, 1983) In turn, the soil environment affects thecomposition of the microbiological population both qualitatively and quantitatively The rate of decom-position of an organic waste depends primarily upon its chemical composition and upon those factorsthat affect the soil environment Factors having the greatest effect on microbial growth and activity willhave the greatest potential for altering the rate of residue decomposition in soil

The ability of the upper 6 in of soil to absorb nutrients and hold water depends upon its physicaland chemical properties of texture, infiltration and permeability, water-holding capacity, bulk density,organic matter content, cation exchange capacity, macronutrient content, salinity, and micronutrientcontent (Hornick, 1983) A typical mineral soil is composed of approximately 45% mineral material(varying proportions of sand, silt, and clay), 25% air and 25% water (i.e., 50% pore space, usually halfsaturated with water), and 5% organic matter, although this is highly variable Any significant change

in the balance of these components could affect the physical and chemical properties of the soil Thismay alter the ability of the soil to support the chemical and biological reactions necessary to degrade,detoxify, inactivate, or immobilize toxic waste constituents

Most soils have a tremendous capacity to detoxify organic chemical wastes by diluting the compounds,acting as a buffering system, and decomposing the material through microbial activity (JRB Associates,Inc., 1984) The most important soil characteristics for this detoxification are those that affect watermovement and contaminant mobility, i.e., infiltration and permeability Certain waste characteristics canalso affect soil infiltration and permeability, and this interaction should be taken into account Table 5.1

lists the site/soil properties that should be identified to be able to predict potential migration of thecontaminating material and indicate what will be necessary for manipulating the soil characteristics foroptimum results Some of the soil factors, however, can be managed only near the surface for enhancingthe soil treatment

Unless all the proper conditions are met for a given compound, biodegradation is not likely to occur(Bitton and Gerba, 1985) Before in situ biological remedial actions can be initiated for treating hazardous

waste–contaminated soils, both the site and waste characteristics must be evaluated (Solanas, Pares,Bayona, and Albaiges, 1984) These features will help determine whether or not a biological approach

is the most feasible treatment option and, if selected, how biodegradation can be used most effectivelywith the prevailing conditions

There are more than 1000 different soil types in the U.S alone (Federle, Dobbins, Thornton-Manning,and Jones, 1986) The U.S Soil Conservation Service has characterized certain chemical and physicalparameters for many of them while preparing soil maps These data are readily available They wouldhelp in predicting biomass and activity in various profiles

The most important soil factors that affect degradation are water; temperature; soil pH; aeration oroxygen supply; available nutrients, i.e., nitrogen (N), phosphorus (P), potassium (K), sulfur (S); oxida-tion/reduction potential; and soil texture and structure Any treatments applied to the soil to enhancecontaminant removal processes must not alter the physical or chemical environment to the extent thatthey would severely restrict microbial growth or biochemical activity (Sims and Bass, 1984) In general,this means that the soil water potential should be greater than –15 bar (Sommers, Gilmore, Wildung,and Beck, 1981); the pH should be between 5 and 9 (Atlas and Bartha, 1981; Sommers, Gilmore,Wildung, and Beck, 1981); and the oxidation-reduction (redox) potential should be between pe + pH of17.5 to 2.7 (Baas Becking, Kaplan, and Moore, 1960) Soil pH and redox boundaries should be carefullymonitored when chemical and biological treatments are combined

Since the activity of microorganisms is so dependent upon soil conditions, modification of soilproperties is a viable method of enhancing the microbial activity in the soil (Sims and Bass, 1984) Tovary these factors for use as a treatment technology, the following information is required:

Characterization and concentration of wastes, both organics and inorganics, at the site;

Microorganisms present at site;

Biodegradability of waste constituents (half-life, rate constant);

Biodegradation products, particularly hazardous products;

Depth, profile, and areal distribution of constituents;

Soil moisture;

Other soil properties for biological activity (pH, Eh, oxygen content, nutrient content, organic matter,temperature);

Trafficability of soil and site

Site location/topography and slope Hydraulic properties and conditions

Amount and type of coarse fragments Depth to impermeable layer or bedrock

a Factors that may be managed to enhance soil treatment with shallow depth

b Factors that may be managed to enhance soil treatment

Source: JRB Associates, Inc Report prepared for Municipal Environmental Research Laboratory, Cincinnati, OH, 1984.

PB 85-124899.

The influence of soil factors, such as temperature and nutrient concentration, on phenol mineralization,for example, shows great variability as a function of soil type and horizon (Thornton-Manning, Jones,and Federle, 1987) Most of these factors do not function independently; i.e., a change in one may effect

a change in others (Parr, Sikora, and Burge, 1983) While the soil factors play an important role inbiodegradation, because of these interactions, it is not always easy to predict a priori how temperature

or another environmental variable will affect biodegradation in a given soil environment Manning, Jones, and Federle, 1987) However, if any of the factors that affect degradation processes insoil are at less than an optimum level, microbial activity will be lowered accordingly and substratedecomposition decreased (Parr, Sikora, and Burge, 1983)

(Thornton-The inherent capacity of soil to degrade toxicants by chemical and biological mechanisms can bemaximized by identification of the soil conditions that promote the degradation of each toxicant andmanipulation of the soil environment to bring about these conditions (Arthur D Little, Inc., 1976).Although each toxicant, in general, has a unique set of ideal soil conditions for degradation, for somecompounds these ideal conditions overlap, and more than one toxic substance can be the focus of soilmanipulation at one time For other compounds, the ideal conditions do not overlap and are sometimeseven contradictory; these materials must be treated in series

Table 5.2 lists the soil factors that may have to be modified during the use of various treatmenttechnologies (Sims and Bass, 1984)

Biodegradation of waste chemicals in the soil requires water for microbial growth and for diffusion ofnutrients and by-products during the breakdown process (JRB Associates, Inc., 1984) Extremes of verywet or very dry soil moisture markedly reduce waste biodegradation rates (Arora, Cantor, and Nemeth,1982) Aerobic waste hydrocarbon decomposition is diminished under saturated soil moisture conditionsbecause of low oxygen supply, while under very dry conditions, microbial activity is hindered due toinsufficient moisture levels necessary for microbial metabolism (CONCAWE, 1980)

A typical soil is about 50% pore space and 50% solid matter (JRB Associates, Inc., 1984) Waterentering the soil fills the pore spaces until they are full The water then continues to move down into thesubsoil, displacing air as it goes The soil is saturated when it is at its maximum retentive capacity Whenwater then drains from the pores, the soil becomes unsaturated Soils with large pores, such as sands, losewater rapidly Larger pores are a less hospitable environment for microorganisms (Turco and Sadowsky,1995), whereas the smaller pores inside the aggregate retain water (Papendick and Campbell, 1981)

If the soil is too impermeable, it will be difficult to circulate treatment agents or to withdraw thepolluted water (Nielsen, 1983) Soils with a mixture of pore sizes, such as loamy soils, hold more water

at saturation and lose water more slowly The density and texture of the soil determine the water-holdingcapacity, which in turn affects the available oxygen, redox potential, and microbial activity (Parr, Sikora,and Burge, 1983) The actual microbial species composition of a soil is often dependent upon wateravailability The migration of organisms in the soil can also be affected by pore size Small bacteria are

on the order of 0.5 to 1.0 µm in diameter (Bitton and Gerba, 1985) Larger bacteria tend to be immobilized

in soils by physical straining or filtering

The water content of soil typically ranges from 15 to 35 vol% (Huddleston, Bleckmann, and Wolfe,1986) At 35%, most soils are water saturated At the other extreme, the concentration can drop lowerthan 15% under unusually arid conditions Soil water content is commonly addressed as percent of soilwater-holding capacity A soil water-holding capacity range of 25 to 100% is typically equivalent to arange of 7 to 28% volume percent Dibble and Bartha (1979) report optimal biodegradation at a soilwater-holding capacity of 30 to 90%

Table 5.3 shows some of the conditions that can be selectively altered for removal of anthropogeniccompounds by particular groups of microorganisms

Field capacity refers to the percentage of water remaining in a soil after having been saturated andfree gravitational drainage has ceased (JRB Associates, Inc., 1984) Gravitational water movement isimportant for mobilizing contaminants and nutrients, due to leaching Slow drainage can reduce microbialactivity as a result of poor aeration, change in oxidation-reduction potential, change in nutrient status,and increased concentration of natural minerals or contaminants to toxic levels in the pore water Theamount of water held in a soil between field capacity and the permanent wilting point for plants is known

as available water This is the water available for plants and for soil microbial and chemical reactions

Bacterial activity is highest in the presence of moisture (JRB Associates, Inc., 1984) Several authorshave indicated ranges of moisture for optimum biodegradation (Bossert, Kachel, and Bartha, 1984; Ryan,Hanson, and Loehr, 1986; Huddleston, Bleckman, and Wolfe, 1986) Some indicate that 30 to 90% offield capacity is needed Others, that 50 to 80% is a better range Based on first-order regressionrelationships for O2 uptake rates, moisture addition of 35 to 50% field capacity was found to accelerate

in situ respiration in a JP-4-contaminated soil (Dupont, Doucette, and Hinchee, 1991) The aerobic

biodegradation of simple or complex organic material in soil is commonly greatest at 50 to 70% of thesoil water-holding (field) capacity (Pramer and Bartha, 1972) Inhibition at levels below 30 to 40% isdue to inadequate water activity, and high values interfere with soil aeration The dependency on soilwater content for biodegradation of petroleum constituents is compound specific and probably also soilspecific (Holman and Tsang, 1995) Moisture is a critical parameter for degradation of two-, three-, andfour-ring polycyclic aromatic hydrocarbons (PAHs), and it has been found that degradation is consider-ably greater at 80% than at 40% of field capacity (Loehr, 1992).

Holman and Tsang (1995) determined that a water content of 50 to 70% of field capacity was optimumfor biodegradation of aromatic hydrocarbons to proceed at maximum rate For simple monoaromaticand diaromatic hydrocarbons, such as toluene and naphthalene, a first-order kinetic model provides agood fit to mineralization data over a range of soil moisture content However, for larger PAHs, such asphenanthrene and anthracene, the model provides a good fit only at soil water content below 50% Sincelong-chain aliphatic hydrocarbons have such a low solubility, their mineralization is little affected bythe soil water content

Fungi — Yeast, mold pH < 5, ae-mae; high O2 tension,

pH < 5 moisture about 50%

Attacks and partially degrades compounds not readily metabolized by other organisms; wide range of nonspecific enzymes

carbon flux

Self-sustaining population, light is primary energy source, partially degrades certain complex compounds, photochemical reactions, oxygenates effluent, no aeration needed, supports growth of other microbes, effective in bioaccumulation of hydrophobic substances

Cyanobacteria

(blue-green algae)

ae-mae, an; light: 600 to 700 nm;

low carbon flux

See algae Bacteria

Heterotrophs (aerobic) ae; proper organic substrate,

growth factors as required;

Eh: 0.45 to 0.2 V

For many compounds degradation is more complete and faster than under anaerobic conditions, high sludge production

Anaerobic (fastidious) an; Eh: <-0.2 to –0.4 V Conditions for abiotic or biological reductive

dechlorination, certain detoxification reactions not possible under aerobic conditions; no aeration, little sludge produced

Facultative anaerobes ae, mae-an; Eh: <-0.2 V No aeration, reductive dechlorination possible Photosynthetic bacteria

Purple sulfur an (light), mae (dark); Eh: 0 to

–0.2 V; S -2 : 2 to 8 mM, 0.4 to

1 mM; light: 800 to 890 nm at

1000 to 2000 lux, high intensities near limit; low C flux

Self-sustaining population able to use light energy, conditions right for reductive dechlorination, no aeration

Purple nonsulfur an; Eh: 0 to –0.2 V; light: 800 to

890 nm; low C flux

See purple sulfur bacteria, also nonspecific enzymes

23 to 28 o C, urea as nitrogen source

Universal scavengers with range of complex organic substrates often not used by other microbes Oligotrophs (from almost

any group above)

ae; carbon flux of <1 mg/L/d;

favorable attachment sites

Removal of organic contaminants in trace concentrations, many inducible enzymes for multiple substrates

Abbreviations: ae = aerobic; mae = microaerophilic (<0.2 atm oxygen); an = anaerobic.

a Possible characteristics for selection, not growth range.

Source: From Kobayashi, H and Rittmann, B.E Environ Sci Technol. 16:170A–183A American Chemical Society Washington, D.C., 1982 With permission.

There is a dramatic difference in characteristics of microbial communities as a result of differentwater content, which parallels mineralization measurements (Holman and Tsang, 1995) The greatestdiversity and activity of microorganisms and the highest population densities are consistently observed

in the sandy, water-bearing strata, whereas the dense, dry-clay layer zones have the least microbiologicalactivity (Fredrickson and Hicks, 1987)

Soil gas humidities <30% cause considerable retardation of hydrocarbon vapors in all media terman, Kulshrestha, and Cheng, 1995) Retardation coefficients decrease but remain large with increas-ing humidity in organic-rich soils Based on soil–water isotherms, there may be competitive sorptionbetween hydrocarbon and water vapors on soil surfaces, especially the mineral fraction

(Bat-Where it is necessary to predict and interpret the response of microorganisms in soils to organicwastes, both the water content and water potential should be reported (Parr, Sikora, and Burge, 1983).Water potential is useful for quantifying the energy status of water in soils containing waste chemicals.Generally, with decreasing water potentials, fewer organisms are able to grow and reproduce; andbacterial activity is usually greatest at high water potentials (wet conditions) Species composition ofthe soil microflora is regulated largely by water availability, which, in turn, is governed essentially bythe energy of the water in contact with the soil or waste

Some fungi can tolerate dry soils but do not grow well if the soil is wet (Clark, 1967) Bacteria may

be antagonistic to fungi under moister conditions At low potentials, bacteria are less active, allowingfungi to predominate (Cook and Papendick, 1970) Microbial decomposition of organic material in driersoils is probably due primarily to fungi (Gray, 1978; Harris, 1981) When soil becomes too dry, manymicroorganisms form spores, cysts, or other resistant forms, while many others are killed by desiccation(JRB Associates, Inc., 1984)

A well-drained soil (e.g., a loamy soil) is one in which water is removed readily but not rapidly (JRBAssociates, Inc., 1984); a poorly drained soil (e.g., a poorly structured fine soil) remains waterloggedfor extended periods of time, producing reducing conditions and insufficient oxygen for biologicalactivity; and an excessively drained soil (e.g., a sandy soil) is one in which water can be removed readily

to the point that drought conditions occur For in situ treatment of hazardous waste–contaminated soils,the most desirable soil would be one in which permeability is only large enough to maximize soilattenuation processes (e.g., adequate aeration for aerobic microbial degradation) while still minimizingleaching Although fine-textured soils may have the maximum total water-holding capacity, medium-textured soils have the maximum available water due to favorable pore size distribution

Control of moisture content of soils at an in situ treatment site may be essential for control andoptimization of some degradative and sorptive processes, as well as for suppression of volatilization ofsome hazardous constituents (Sims and Bass, 1984) The moisture content of soil may be controlled toimmobilize constituents in contaminated soils and to allow additional time for accomplishing biologicaldegradation When contaminants are immobilized by this technique and anaerobic decomposition isdesired, anaerobiosis must be achieved by a means other than flooding, such as soil compaction ororganic matter addition Control of soil moisture may be achieved through irrigation, drainage, or acombination of methods

The need for moisture was demonstrated in the efforts to remediate oil-polluted Kuwaiti desert soil(Radwan, Sorkhoh, Fardoun, and Al-Hasan, 1995) The amount of alkanes in the untreated controlsremained constant during the dry hot months then decreased during the rainy season After 1 year, thedesert had cleaned itself of half the contaminating extractable alkanes, but had required moisture to do

so Fertilized soils reduced these compounds to about a third in that time

In another instance, a subsurface drip irrigation system was used to increase soil moisture duringbioventing dry, sandy soils contaminated with gasoline, JP-5 jet fuel, and diesel fuel to a depth of 24 m(Zwick, Leeson, Hinchee, Hoeppel, and Bowling, 1995) In situ respiration rates increased significantly

as a result

Sometimes a site with shallow depth contamination may require soil mixing to dilute the wastes andincorporate nutrients and oxygen, as well as to enhance soil drying (Sims and Bass, 1984) It may benecessary to install a drainage system to reduce soil moisture Increasing soil temperature will enhancesurface soil drying This can be achieved with landfarming However, drying the soil may retard microbialactivity, as well as increase volatilization of volatile waste components

Excess moisture, extremely dry conditions, pooling, or flooding should be avoided (Zitrides, 1983).Biodecontamination programs should not be conducted during heavy rains or drought However, an

observed lack of inhibition at 30% of the field capacity suggests that the moisture requirement formaximum activity on hydrophobic petroleum may be different than the optimal moisture levels for thebiodegradation of hydrophilic substrates (Dibble and Bartha, 1979a).

Rainfall dissolves contaminants and acts as a carrier as it percolates through the soil on its way tothe groundwater, which can be useful to the bioremediation plan, if this is desired (Dietz, 1980).Rainwater also keeps the contaminated soil moist, and microorganisms will utilize the oxygen dissolved

in interstitial water droplets (Thibault and Elliott, 1980)

Many organisms are capable of metabolic activity at water potentials lower than –15 bar (Soil ScienceSociety of America, 1981) The lower limit for all bacterial activity is probably about –80 bar, but someorganisms cease activities at –5 bar Although many microbial functions continue in soils at –15 bar ordrier, optimum biochemical activity is usually observed at soil water potentials of –0.1 to 1.0 bar(Sommers, Gilmore, Wildung, and Beck, 1981) The kinds of microorganisms that are metabolicallyactive in the soil will be affected Degradation rates are highest at soil water potential between 0 and–1 bar When natural precipitation cannot maintain near optimal soil moisture for microbial activity,irrigation may be necessary (Sims and Bass, 1984)

Moisture control is widely practiced in agriculture; however, there is little information on its use tostimulate biological degradation of hazardous materials in soil (Sims and Bass, 1984) Most laboratorystudies have been conducted at or near optimal soil moisture The success of this technology dependsupon the biodegradability of the waste constituents and the suitability of the site and soil for moisturecontrol Although degradation of hazardous organic compounds may be accelerated by soil moistureoptimization, effectiveness of this treatment approach may be enhanced by combination with othertechniques to increase biological activity The technology is reliable in that it has been used in agriculture,but retreatment is necessary There may be problems with leaching of soluble hazardous compoundsand erosion

Control of soil moisture content can be practiced to optimize degradative and sorptive processes andmay be achieved by several means (Sims and Bass, 1984) Supplemental water may be added to the site(irrigation), excess water may be removed (drainage, well points), or these methods can be combinedwith other techniques, such as using soil additives, for greater moisture control

Soil may be irrigated by subirrigation, surface irrigation, or overhead (sprinkler) irrigation (Fry andGrey, 1971) With subirrigation, water is applied below the ground surface and moves upward by capillaryaction Water with high salinity may allow accumulation of salts in the surface soil, with an adverseeffect on microbial activity The site must be nearly level and smooth, with either a natural or perchedwater table, which can be maintained at a desired elevation Check dams and gates in open ditches orjointed perforated pipe can be used to maintain the water level in the soil These systems may be limited

by the restrictive site criteria A subirrigation system might be combined with a drainage system tooptimize soil moisture content At a hazardous waste site, though, raising the water table might produceundesirable groundwater contamination

With trickle irrigation, filtered water is supplied directly on or below the soil surface through anextensive pipe network with low-flow-rate outlets only to areas that require irrigation (Fry and Grey,1971) Coverage of an area will not be uniform, but with proper management, percolation and evaporationlosses can be reduced For most in-place treatment sites, this method would probably not be appropriate,but it may be applicable in an area where only “hot spots” of wastes are being treated

Surface irrigation includes flood, furrow, or corrugation irrigation (Fry and Grey, 1971) Since site migration of hazardous constituents to groundwaters or surface waters should normally be prevented,surface irrigation should be considered with caution Contaminated water may also be a hazard to on-site personnel

off-In flood irrigation, water covers the surface of a soil in a continuous sheet (Fry and Grey, 1971).Theoretically, water should remain in place just long enough to apply the desired amount, but this isdifficult or impossible to achieve under field conditions Widrig and Manning (1995) determined thatcontinuous saturation by flooding with nitrogen and phosphorus amendments was not as effective asperiodic operation, consisting of flooding with nutrients, followed by draining and forced aeration.Monitoring CO2 and O2 levels in situ may allow optimization of the timing of flooding and aerationevents to increase degradation rates

In furrow irrigation, water is applied in narrow channels or furrows As the water runs down thefurrow, part of it infiltrates the soil (Fry and Grey, 1971) Irrigation of the soil between furrows requiresconsiderable lateral water movement Salts may accumulate between furrows Furrow irrigationfrequently requires extensive land preparation, which usually would not be possible or desirable at ahazardous waste site because of contamination and safety considerations.

In corrugation irrigation, as with furrow irrigation, water is applied in small furrows from a headditch (Fry and Grey, 1971) The furrows are used in this case only to guide the water, and overflooding

of the furrows can occur

In general, control and uniform application of water is difficult with surface irrigation Also, soilshigh in clay content tend to seal when water floods the surface, limiting water infiltration

The basic sprinkler irrigation system consists of a pump to transfer water from the source to the site,

a pipe or pipes leading from the pump to the sprinkler heads, and the spray nozzles (Fry and Grey,1971) Sprinkler irrigation has many advantages For instance, application rates can be adjusted for soils

of different textures, even within the same area; water can be distributed more uniformly; and erosionand runoff of irrigation water can be controlled or eliminated Sprinkler irrigation is also possible onsteep, sloping land and irregular terrain This method usually requires less water than surface flooding,and the amount of water applied can be controlled to meet the needs of the in-place treatment technique.Also, a larger soil surface area can be covered, which could facilitate soil washing

There are several types of sprinkler irrigation systems (Fry and Grey, 1971):

1 Permanent installations with buried main and lateral lines;

2 Semipermanent systems with fixed main lines and portable laterals;

3 Fully portable systems with portable main lines and laterals, as well as a portable pumping plant

The first two types (especially the first) would probably not be cost-effective or appropriate for ahazardous waste site because of the required land disturbance for installation and the limited time periodfor execution of the treatment

There are fully portable systems available These may have hand-moved or mechanically movedlaterals (Fry and Grey, 1971) Portable systems are useful in difficult areas, such as forests, where theywill not interfere with trees Mechanically moved laterals may be side-roll/wheel-move, center-pivotsystems, or traveling sprinklers This equipment is more expensive but requires much less labor thanthe hand-moved systems The health and safety of workers must be considered, as well as the cost, inthe choice of an appropriate system

Subsurface drains can be used to lower the water table, while surface drains are used where subsurfacedrainage is impractical (e.g., impermeable soils, excavation difficult) to remove surface water or lowerthe water table (Donnan and Schwab, 1974) Construction materials for the drainage systems includeclay or concrete tile, corrugated metal pipe, and plastic tubing Selection of the materials depends uponstrength requirements, chemical compatibility, and cost

Various additives are available to enhance moisture control; e.g., the water-retaining capacity of the soilcan be enhanced by adding water-storing substances (Nimah, Ryan, and Chaudhry, 1983) Evaporationretardants are available for retaining soil moisture There are also water-repelling agents for diminishingwater absorption by soils Water-repelling soils can be treated with surface-active wetting agents toimprove water infiltration and percolation Surface-active agents also accelerate soil drainage, modifysoil structure, disperse clays, and make soil more compactable

5.1.2 TEMPERATURE

Soil temperature is one of the more important factors controlling microbiological activity and the rate

of organic matter decomposition (Sims and Bass, 1984) Temperatures of both air and soil affect therate of biological degradation processes in the soil, as well as the soil moisture content (JRB Associates,Inc., 1984) Temperature affects the physical nature and composition of the petroleum, the rate ofmicrobial hydrocarbon metabolism, and the composition of the microbial communities (Atlas, 1994).There is an optimum temperature, beyond which biological activity often decreases rapidly, thus dis-playing a growth curve that is skewed to the right (JRB Associates, Inc., 1984)

Generally, raising the temperature increases the rate of degradation of organic compounds in soil(JRB Associates, Inc., 1982) Microbial growth usually doubles for every 10°C increase (Thibault andElliott, 1979) There is a decrease in adsorption with rising temperature, which makes more organicsavailable for the microorganisms to degrade (JRB Associates, Inc., 1984) On the other hand, highertemperatures increase evaporation of short-chain alkanes and other low-molecular-weight hydrocarbons,which usually cause solvent-type membrane toxicity to microorganisms (Atlas, 1994) They also decreasethe viscosity of the petroleum hydrocarbons and their solubility in the soil aqueous phase High tem-peratures, well above those normally experienced in soil, cause very rapid decreases in growth andmetabolism and become lethal (Huddleston, Bleckmann, and Wolfe, 1986) If temperatures exceed 41

to 42°C, enzymes in the bacteria normally begin to break down, and life processes fail (Lapinskas, 1989).Conversely, a lowering of the temperature is associated with a slowing of the microbial growth rate(Thibault and Elliott, 1979) Low temperatures can lengthen the acclimation period and delay onset ofbiodegradation (Zhou and Crawford, 1995) A microbial community will undergo an adaptation orselection process in the mineralization of a compound, which is reflected in a lag period that oftenincreases with decreasing temperature (Thornton-Manning, Jones, and Federle, 1987) Low temperaturesalso can decrease microbial enzymatic activity — i.e., the “Q10” effect (Zhou and Crawford, 1995) Lowtemperatures are not lethal to microorganisms, although repeated freezing and thawing will rupture some(Huddleston, Bleckmann, and Wolfe, 1986)

Microbial utilization of hydrocarbons can occur at temperatures ranging from –2 to 70°C (TexasResearch Institute, Inc., 1982) Most soils, especially those in cold climates, contain psychrophilicmicroorganisms that grow best at temperatures below 20°C (JRB Associates, Inc., 1984) and are effective

at temperatures below 0°C Biodegradation can take place at a temperature of 5°C, but hydrocarbonsare degraded more slowly at lower temperatures (Parr, Sikora, and Burge, 1983) Walworth and Reynolds(1995) report that bioremediation is effective for treating petroleum-contaminated soils in cold areas;however, diesel fuel loss is certainly greater in soil at 20 than at 10°C At 10°C, the bioremediation ratesare not affected by addition of phosphorus or nitrogen, but they are increased at 20°C by addition ofphosphorus but not nitrogen Dibble and Bartha (1979) found the optimum temperature for biodegrada-tion to be 20°C or higher

Whyte, Greer, and Inniss (1996) tested 135 psychrotrophic microorganisms for the ability to alize petroleum hydrocarbons A number of strains mineralized toluene, naphthalene, dodecane, andhexadecane Rhodococcus sp Q15 was able to mineralize the C28n-paraffin, octacosane All the psy-chrotrophic biodegradative isolates were capable of mineralization activity at both 23 and 5°C, indicatingtheir potential for low-temperature bioremediation of petroleum hydrocarbon–contaminated sites.Soils in hot environments usually support many thermophilic microorganisms that are effective attemperatures above 60°C (Texas Research Institute, Inc., 1982) However, most soil microorganisms aremesophiles and exhibit maximum growth in the range of 20 to 35°C (Parr, Sikora, and Burge, 1983).The majority of hydrocarbon utilizers are most active in this range Since many organisms multiply well

miner-at laborminer-atory temperminer-atures of 25 to 37°C but not miner-at lower environmental temperminer-atures, it would bebeneficial to isolate appropriate organisms at temperatures and in media that correspond to the charac-teristics of the contaminated site (Alexander, 1994)

Temperatures in the thermophilic range (50 to 60°C) were shown to greatly accelerate decomposition

of organic matter, in general (Parr, Sikora, and Burge, 1983) At these temperatures, actinomycetes will

be naturally predominant over fungi and bacteria Therefore, in certain situations, composting may offerpotential for maximizing the biodegradation rate of waste industrial chemicals It should be noted, however,that in another investigation in a test treatment facility, it was found that several aromatic hydrocarbonswere not metabolized at 55°C, but were metabolized at 30°C (Phillips and Brown, 1975), while otherresearchers reported a leveling-off of the hydrocarbon biodegradation rate in soil above 20°C (Dibble and

Bartha, 1979a) Although elevated temperature has some advantage for potentially limiting the development

of pathogenic microorganisms, too high a temperature would not be beneficial for stimulating petroleumbiodegradation (Phillips and Brown, 1975) The increased availability of more-toxic hydrocarbons at highertemperatures may counteract the stimulation of metabolic processes (Dibble and Bartha, 1979a).Mutant organisms are being developed to provide the optimal degradation at any given temperature Acommercially available mutant bacterial formulation (PETROBAC® Mutant Bacterial HydrocarbonDegrader) provides degradation of crude oil over a range of temperatures from 5 to 35°C, with the greatestamount of degradation in the shortest amount of time at the higher temperatures (Thibault and Elliott, 1979)

A temperature gradient exists in the soil (Ahlert and Kosson, 1983) As a result of heat transferphenomena, temperature responds less to daily weather fluctuations at increased depths Microorganismsnear the surface of the soil column must adapt more readily to temperature fluctuations than those atgreater depth Thus, the seasonal and geographic variations play a role in degradation rates (JRBAssociates, Inc., 1984) Disposal sites for oil can be chosen in warm areas that receive direct sunlight

to assure temperatures suitable for rapid metabolism by mesophilic microorganisms (Atlas, 1977) Even

in near-Arctic environments, absorbance of solar energy raises temperatures into a range that allows formesophilic microbial oil degradation (Atlas and Schofield, 1975)

Soil temperature is difficult to control in a field situation, but can be modified by regulating theincoming and outgoing radiation, or by changing the thermal properties of the soil (Baver, Gardner, andGardner, 1972) Vegetation plays a significant role in soil temperature because of the insulating properties

of plant cover (Sims and Bass, 1984) Bare soil unprotected from the direct rays from the sun becomesvery warm during the hottest part of the day, but also loses its heat rapidly at night and during colderseasons In the winter, vegetation acts as an insulator to reduce heat lost from the soil Frost penetration

is more rapid and deeper under bare soils than under a vegetative cover On the other hand, during thesummer months, a well-vegetated soil does not become as warm as a bare soil These fluctuations insoil temperature decrease with increasing depth (Thornton-Manning, Jones, and Federle, 1987).Soil temperature can be modified by soil moisture control and by the use of mulches of natural orartificial materials (JRB Associates, Inc., 1984) Mulches can affect soil temperature in several ways Ingeneral, they reduce diurnal and seasonal fluctuations in soil temperature (Sims and Bass, 1984) In themiddle of summer, there is little overall temperature difference between mulched and bare plots, butmulched soil is warmer in spring, winter, and fall, and warms up more slowly in the spring

Mulches with low thermal conductivities decrease heat flow both into and out of the soil; thus, soilwill be cooler during the day and warmer during the night White paper, plastic, or other types of whitemulch increase the reflection of incoming radiation, thereby reducing excessive heating during the day

A transparent plastic mulch transmits solar energy to the soil and produces a greenhouse effect A blackpaper or plastic mulch absorbs radiant energy during the day and reduces heat loss at night Placing ablack covering over the soil to increase the soil temperature during the winter has been suggested as ameans of overcoming the problem of slower biodegradation at the lower winter temperatures (Guidinand Syratt, 1975) Use of polyethylene sheeting as a landfarming cover during treatment of crudeoil–contaminated soil does not appear to affect biodegradation kinetics adversely under laboratoryconditions (Huesemann and Moore, 1993) Humic substances are dark, which increases the heat absorp-tion of the surface soil (Sims and Bass, 1984) Use of film mulch as a means of stimulating waste oilbiodegradation by increasing soil temperatures during the winter, however, would preclude tilling of thesoil and, thus, decrease its aeration (Dibble and Bartha, 1979a) Some researchers believe this wouldnot have an overall beneficial effect and may, in fact, be unnecessary, since the albedo decrease due tooil contamination can raise the temperature in the upper 10 to 20 cm of tundra soils as much as 5°C(Freedman and Hutchinson, 1976)

Mulches are also used to protect soil surfaces from erosion, reduce water and sediment runoff,conserve moisture, prevent surface compaction or crusting, and help establish plant cover (Soil Conser-vation Service, 1979) The type of mulch required determines the application method (Sims and Bass,1984) Commercial machines for spraying mulches are available (Soil Conservation Service, 1979).Hydromulching is a process in which seed, fertilizer, and mulch are applied as a slurry To apply plasticmulches, equipment is towed behind a tractor and mechanically applies plastic strips that are sealed atthe edges with soil For treatment of large areas, special machines that glue polyethylene strips togetherare available (Mulder, 1979) Table 5.4 describes the organic materials available for use as mulch andthe situations when each would be most suitable

Irrigation increases the heat capacity of the soil, lowers air temperature over the soil, raises thehumidity of the air, and increases thermal conductivity, resulting in a reduction of daily soil temperaturevariations (Schweizer, 1976) Sprinkle irrigation, for example, has been used for temperature control,specifically frost protection in winter and cooling in summer and for reduction of soil erosion by wind(Schwab, Frevert, Edminster, and Barhes, 1981) Drainage decreases the heat capacity, which raises thesoil temperature Elimination of excess water in spring causes a more rapid temperature increase Theaddition of humic substances improves soil structure, thus improving soil drainability, resulting indirectly

in an increase in the insulative capacity of the soil

Several physical characteristics of the soil surface can be modified to alter soil temperature (Schweizer,1976) Compaction of the soil surface will increase the density and, thus, the thermal conductivity.Tillage, on the other hand, creates a surface mulch that, when dry, reduces heat flow from the surface

to the subsurface The diurnal temperature variation in a cultivated soil is often much greater than in anuntilled soil A loosened soil has more surface area exposed to the sun but is colder at night and moresusceptible to frost

Raising the temperature of a contaminated zone can also be achieved by pumping in heated water orrecirculating groundwater through a surface heating unit (U.S EPA, 1985a) Tentlike structures can beerected over treatment beds to elevate temperature, especially in winter (Ellis, Harold, and Kronberg, 1991)

Soil pH contributes to the surface charge on many colloidal-sized soil particles (JRB Associates, Inc.,1984) Clays have a permanent negative charge, and it is primarily their coatings of organic andamorphous materials that change in charge Thus, the pH of the groundwater and soil water in the vadose

Small-grain straw or

tame hay

Undamaged, air-dried threshed straw, free of undesirable weed seed

Spread uniformly — at least of ground should be visible

to avoid smothering seeding; anchor either during application or immediately after placement to avoid loss by wind or water; straw anchored in place is excellent on permanent seedings

Corn stalks chopped

excessive coarse material

Excellent around shrubs; may create problems with weeds Wood chips and bark Air-dried, free from objectionable

coarse material

Most effective as mulch around ornamentals, etc.; resistant

to wind blowing; may require anchoring with netting to prevent washing or floating off

seeded areas subject to foot traffic (approx weight —

1 ton/yd 3 ) Wood excelsior mats Blanket of excelsior fibers with a

net backing on one side

Roll 36 in × 30 yd covers 16 yd 2 ; use without additional mulch; tie down as specified by manufacturer

Jute, mesh, or net Woven jute yarn with in

openings

Roll 48 in × 75 yd weighs 90 lb and covers 100 yd 2

Source: Soil Conservation Service Guide for Sediment Control on Construction Sites in North Carolina. U.S Department

of Agriculture, Raleigh, NC, 1979.

1 / 4

1 / 4

3 / 4

zone determines the degree of anion or cation adsorption by soil particles In soils with pH-dependentcharge, lowering the pH decreases the net negative charge and, thus, decreases anion repulsion orincreases anion adsorption (Hornick, 1983) The soil pH may affect the solubility, mobility, and ionizedforms of contaminants (JRB Associates, Inc., 1984).

Biological activity in the soil is greatly affected by the pH, through the availability of nutrients andtoxicants and the tolerance of organisms to pH variations Some microorganisms can survive within awide pH range, while others can tolerate only small variations The optimum pH for rapid decomposition

of wastes and residues is usually in the range of 6.5 to 8.5 Bacteria and actinomycetes have pH optimanear 7.0 A soil pH of 7.8 should be close to the optimum (Dibble and Bartha, 1979a) If the soil isacidic, these organisms often cannot compete effectively with soil fungi for available nutrients The pHcan influence the solubility or availability of macro- (especially phosphorus) and micronutrients, themobility of potentially toxic materials, and the reactivity of minerals (e.g., iron or calcium) (Parr, Sikora,and Burge, 1983)

Hydrocarbon-contaminated soil could contain a number of heavy metals that are potentially toxic tothe environment (Streebin, Robertson, Callender, Doty, and Bagawandoss, 1984) The pH range of 6.5 to8.0 is also optimum for the formation of insoluble precipitates and, thus, results in the immobilization

of certain heavy metals pH is the most important aspect of the reaction between heavy metals and soils(Leeper, 1978) The leaching of metals will not be a problem at treatment sites employing proper pHcontrol (Dibble and Bartha, 1979c)

Contamination by hydrocarbons can change the pH of the soil (Amadi, Abbey, and Nima, 1996).After exposure to oil spillage for 17 years, the pH of the soil varied from 4 to 6 in heavy and moderatelyimpacted zones Soil nutrients were similar in both areas Petroleum hydrocarbon utilizers correlatedpositively with the distribution of oil Aerobic nitrifiers, however, were more abundant in the heavy thanthe moderate zone, while anaerobic nitrifiers were higher in the moderate than in the heavy

Carbonic acid, organic acid intermediates, and nitrate and sulfate (most important for pH < 5) mayaccumulate during aerobic degradation of organic molecules (Zitrides, 1983) This can lower the soil

pH and inhibit biological activity The acid conditions can be controlled with reinoculation or by addition

of lime, which is favorable for the biodegradation of oil (Dibble and Bartha, 1979a)

When acidic wastes in hazardous waste–contaminated soil lower the soil pH, they change themicroorganism distribution (Sims and Bass, 1984; JRB Associates, Inc., 1984) Fungi predominate underacidic conditions (pH < 7) These organisms may transform aromatic hydrocarbons by means of oxyge-nases into arene oxides, the mutagenic forms of PAHs (Baver, Gardner, and Gardner, 1972) Bacteria,

on the other hand, growing better at a neutral or slightly basic pH, would carry out the dioxygenation

of the aromatic nucleus to form a cis-glycol as the first stable intermediate, instead of the arene oxide.Higher organisms (above fungi) do not possess the necessary oxygenases and, thus, form trans aromaticdiols, which tend to polymerize

These differences in the mechanism of aromatic hydrocarbon metabolism by microorganisms haveimportant implications concerning engineering techniques for controlling and possibly detoxifying sim-ple aromatics and PAHs in contaminated soils (Cerniglia, Hebert, Dodge, Szaniszlo, and Gibson, 1979)

It appears that selection for dominance of the microbial community by bacteria may avoid the formation

of mutagens, and that pH may serve as an important engineering tool to direct the pathway of PAHdegradation

Control of soil pH at an in-place hazardous waste treatment site is a critical factor in several treatmenttechniques, including metal immobilization and optimum microbial activity (Sims and Bass, 1984) The

pH of different soil types can vary The goal of soil pH adjustment in agricultural application usually is

to increase the pH to near neutral values, since most natural soils tend to be slightly acidic Areas ofthe country in which the need for increasing soil pH is greatest are the humid regions of the East, South,Middle West, and Northwest States In areas where rainfall is low and leaching is minimal, such as parts

of the Great Plain States and the arid, irrigated saline soils of the Southwest, Intermountain, and FarWest States, pH adjustment is usually not necessary but may require reduction

A calcareous (containing calcium carbonate) soil can range from pH 7 to 8.3 (JRB Associates, Inc.,1984) A sodic (high in sodium carbonate) soil can go as high as pH 8.5 to 10 Saline soils tend to bearound pH 7 The soil pH may need to be lowered by adjusting with sulfur or other acid-forming compounds,

or raised by adding crushed limestone or lime products to bring it between pH 5.5 and 8.5 to encouragemicrobial activity Phosphorus solubility is maximized at pH 6.5; this may be the ideal soil pH

Since it is common to isolate microorganisms that grow well around pH 7, those selected for duction into the environment may not survive or function as desired, if the pH of the contaminated soil isnot in that range (Alexander, 1994) For growth of the appropriate organisms, the isolation medium should

reintro-be maintained at pH values similar to those at the site of concern (Zaidi, Murakami, and Alexander, 1989)

Liming is a frequent agricultural practice and is the most common method of controlling pH, whileacidification is much less common (Sims and Bass, 1984) Methods have been developed to determinethe lime requirement of soils, taking into account the buffering capacity of the soil (McLean, 1982) Alime requirement test may be performed to find the loading rate to use for increasing soil pH However,there are no readily available guidelines for reducing soil pH, and the acidification requirements for aparticular soil have to be determined experimentally in the laboratory, taking into account the bufferingcapacity of the waste Thorough mixing is required in the zone of contamination to change the pH.Runoff and minor controls are necessary to control drainages and erosion of the tilled soil The achievablelevel of treatment is high, depending upon the wastes, site, and soil It may be necessary to repeat theprocess during the treatment

Liming is the addition to the soil of any calcium or calcium- and magnesium-containing compoundcapable of reducing acidity (i.e., raising pH) (Sims and Bass, 1984) Lime correctly refers only to calciumoxide, but is commonly used to refer to calcium hydroxide, calcium carbonate, calcium-magnesiumcarbonate, and calcium silicate slags

There are several benefits of liming to biological activity (Sims and Bass, 1984) Manganese andaluminum are toxic to most plants but are less soluble at higher pH values Phosphates and mostmicroelements necessary for plant growth (except molybdenum) are more available at higher pH.Microbial activity is greater at or near neutral pH, which enhances degradation processes, mineralization,and nitrogen transformations (e.g., nitrogen fixation and nitrification)

The liming material to use depends upon several factors (Sims and Bass, 1984) Calcitic and dolomiticlimestones are the most common However, these must be ground in order to be effective quickly, sincethe velocity of reaction depends upon the surface in contact with the soil The finer they are ground, themore rapidly they react with the soil There will usually be a mixture of fine and coarse particles in afinely ground product This material allows a rapid pH change, is relatively long lasting, and is reasonablypriced Many states require that 75 to 100% of the limestone pass an 8- to 10-mesh sieve and that 20 to80% pass anywhere from an 8- to 100-mesh sieve Calcium oxide and calcium hydroxide are manufac-tured as powders and react quickly Other factors to consider in the selection of a limestone areneutralizing value, magnesium content, and cost per ton applied to the land

Lime requirement for soil pH adjustment depends on soil factors, such as soil texture, type of clay,organic matter content, and exchangeable aluminum (Follett, Murphy, and Donahue, 1981) The bufferingcapacity reflects the soil cation exchange capacity and will directly affect the amount of lime needed toadjust soil pH The amount of lime required is also a function of the volume of soil to be treated Theamount of lime necessary in a particular site/soil/waste system can be determined by a commercial soiltesting laboratory in short-term treatability studies or soil-buffer tests (McLean, 1982) Lime requirementsare also affected by acid-forming fertilizers Commonly used liming materials are summarized in Table 5.5.Limestone must be placed where needed, since it does not migrate easily in the soil and is onlyslightly soluble (Sims and Bass, 1984) Therefore, plowing or disking surface-applied lime into the soilmay be required The application of fluid lime is becoming more popular, especially when mixed withfluid nitrogen fertilizer This combination results in fewer passes over the soil, and the lime is available

to counteract acidity produced by the nitrogen Also, limestone has been applied successfully to apharmaceutical wastewater landtreatment facility through a spray irrigation system

The addition of basic waste to acidic soil increases the pH of the surface layer (4 to 18 in.) but notthe subsoil (Brown, 1975) The reaction neutralizes the buffer capacity of the soil Basic waste can causephysical damage to the soil system; however, weak organic bases added to the soil may increase the soilbuffer capacity and exchange capacity as the bases are degraded

Ferrous sulfate can be added to the soil to decrease alkalinity (Arthur D Little, 1976) Under acidicconditions in soils, solubilities of complexed cations, such as those of copper and zinc, increase, and

those of simple ions of iron, manganese, and copper are easily reduced to more soluble forms (JRBAssociates, Inc., 1984) Acidic wastes may also be used as a treatment process for saline-sodic soils.

Although some xenobiotic organic compounds appear to require the slow anaerobic metabolism fordecomposition, most of these compounds are susceptible to attack by aerobic organisms (Alexander,1977; Brunner and Focht, 1983; Jain and Sayler, 1987) Therefore, assuring the aerobiosis of the soilwill enhance the rate of biological decomposition for many compounds

For degradation to occur, microorganisms must utilize an electron acceptor, such as oxygen (Turcoand Sadowsky, 1995) Most biodegradation of petroleum hydrocarbons is aerobic, since hydrocarbonoxidation processes generally require oxygenases (Atlas and Bartha, 1987)

Molecular oxygen, which is soluble in oils, penetrates oil-contaminated soils and sediments to adegree that depends upon depth, the concentration of oil, and the presence of cracks and fissures in soils

or of burrowing worms in sediments (Lee, 1977; Gordon, Dale, and Keizer, 1978) Typically, a gradientoccurs in which biodegradation shows a strong negative relation to depth

The degree to which the soil pore space is filled with water affects the exchange of gases throughthe soil (JRB Associates, Inc., 1984) Microbial respiration, plant root respiration, and the respiration

of other organisms removes oxygen from the soil and replaces it with carbon dioxide Gases diffuse intothe soil from the air above, and gases in the soil diffuse into the air However, the oxygen concentration

in the surface, unsaturated soil may be only half that in air, while carbon dioxide concentrations may

be many times that of air (Brady, 1974)

Air contains 79% nitrogen, which is essentially useless in bioremediation except in removing solved CO2 to aid in pH control (Bergman, Greene, and Davis, 1994) However, the pH can be controlledchemically As a simple diluent, nitrogen also reduces the amount of oxygen that can be dissolved in abody of waste by a factor of five, which decreases the oxygen dissolution rate This in turn may limitthe rate of biodegradation of the contaminant and make it more difficult to control dissolved oxygenconcentration

dis-As soil becomes saturated, the diffusion of gases through the soil is severely restricted In saturatedsoil, oxygen can be consumed faster than it can be replaced, and the soil becomes anaerobic (JRBAssociates, Inc., 1984) This drastically alters the composition of the microflora Facultative anaerobes,which use alternative electron acceptors, such as nitrate (denitrifiers) and strict anaerobic organisms

Calcium Carbonate

Limestone, calcitic CaCO3, 100% purity 100 Neutralization value usually between 90 and 98%

because of impurities; pulverized to desired fineness Limestone, dolomitic 65% CaCO3 + 20%

MgCO3, 87% purity

89 Pure dolomite (50% MgCO3 and 50% CaCO3) has

neutralizing value of 109%; pulverized to desired fineness

Limestone, unslaked

lime, burned lime,

quick lime

depends on purity of raw materials; white powder, difficult to handle — caustic; quick acting; must be mixed with soil or will harden and cake

Hydrated lime, slaked

lime, builder’s lime

Ca(OH)2, 85% purity 85 Prepared by hydrating CaO; white powder, caustic,

difficult to handle; quick acting

earth, and usually quite moist

contains magnesium Waste lime products Extremely variable in

composition

Source: Follett, R.H et al Fertilizers and Soil Amendments. Prentice-Hall, Englewood Cliffs, NJ, 1981 With permission.

become the dominant species While many soil bacteria can grow under anaerobic conditions, thoughless actively, most fungi and actinomycetes do not grow at all (Parr, Sikora, and Burge, 1983) Microbialmetabolism shifts from oxidative to fermentative and becomes less efficient in terms of biosyntheticenergy production (JRB Associates, Inc., 1984) Soil structure and texture primarily determine the size

of soil pores, and hence the water content at which gas diffusion is significantly limited in a given soil,and the rate at which anaerobiosis sets in

Anaerobic reactions are accompanied by the production of malodorous compounds, such as amines,mercaptans, and H2S (Parr, Sikora, and Burge, 1983) These can be phytotoxic, and, if the soil is heavilyoverloaded, it may remain anaerobic for some time However, if the oxygen balance is maintained,relative to the amount of contaminants and the soil conditions, rapid aerobic decomposition will occur,and the end products will be inorganic carbon, nitrogen, and sulfur compounds

The diffusion of air into soil is generally proportional to the square of the air-filled porosity (Turcoand Sadowsky, 1995) Since, the active microorganisms appear to be located in areas with a pore neckdiameter of 6 µm or less, the total exchange area for gases is limited by the pore neck, to an area of up

to 28 µm2

If oil is also present, it can act as a diffusion barrier for oxygen moving into water (Downing andTruesdale, 1955) The diffusivity of oxygen moving in oil is about 2 × 10–3 cm2/s (Schwarzenbach,Gschwend, and Imboden, 1993) If the films are thicker than 100 µm, they can reduce the overall transfervelocity by as much as half Thus, the ability to transfer oxygen will be limited in soils with largeamounts of oil

A great deal of oxygen-containing water is needed in fine-textured subsurface materials (Wilson,Leach, Henson, and Jones, 1986) Biodegradation of most organic contaminants requires approximatelytwo parts of oxygen to completely metabolize one part of organic compound The complete oxidation

of 1 mg of hydrocarbon to carbon dioxide and water requires 3 to 4 mg of oxygen (Texas ResearchInstitute, Inc., 1982) Less oxygen is needed when microbial biomass (new microorganisms) is generated

or when oxidation is not complete Lund and Gudehus (1990) report that 1.0 kg of hydrocarbons requiresabout 1.5 to 3.5 kg oxygen, depending upon the kind of hydrocarbons and the portion of which isconverted into biomass Typically, about half the carbon in hydrocarbons is converted into biomass(Green, Lee, and Jones, 1981) In general, the more oxygen, the faster the biodegradation (Zhou andCrawford, 1995) The concentration of the contaminants is an important factor (Wilson, Leach, Henson,and Jones, 1986) Dissolved oxygen should be maintained above the critical concentration for thepromotion of aerobic activity, which ranges from 0.2 to 2.0 mg/L, with the most common being 0.5 mg/L(U.S EPA, 1985a) Zhou and Crawford (1995) found that the optimal oxygen concentration for gasoline-degrading microorganisms was only about 10%, which was surprisingly low This is about half of theatmospheric oxygen concentration Other authors report that an oxygen concentration of 5% can belimiting to biodegradation (Wuerdemann, Wittmaier, Rinkel, and Hanert, 1994)

The problem is providing the necessary amount of oxygen to the site where it will be used Oxygencan be provided to the subsurface through the use of air, pure oxygen, hydrogen peroxide, or ozone(U.S EPA, 1985a) Oxygen levels can be increased about fivefold by sparging injection wells withoxygen instead of air (Wilson, Leach, Henson, and Jones, 1986) Fluid and semisolid systems can beaerated by means of pumps, propellers, stirrers, spargers, sprayers, and cascades (Texas ResearchInstitute, Inc., 1982) The advantages and disadvantages of various oxygen supply alternatives aresummarized in Table 5.6 (U.S EPA, 1985a)

The flow of oxygen into the system is controlled by oxygen concentration in the carrier and thepermeability of the geological material to that carrier (Wilson, Leach, Henson, and Jones, 1986) Theamount of oxygen available for biodegradation of hydrocarbons varies considerably with the particularelectron acceptor and its carrier medium (Hinchee and Miller, 1990) With water as a carrier, oxygen inair will supply 8.0 mg O2/L water, and 400,000 kg water would be required to degrade 1 kg hydrocarbon.Pure oxygen delivers 40.0 mg O2 mg /L water and requires 80,000 kg water/kg hydrocarbon If theoxygen comes from H2O2, a level of 100.0 mg O2/L H2O2 in water would require 65,000 kg carrier/kghydrocarbon, or a level of 500.0 mg/L H2O2 in water would require 13,000 kg/kg hydrocarbon With20.9% oxygen in air, 13 kg/kg hydrocarbon would be needed Dupont, Doucette, and Hinchee (1991)report that to obtain 1 g of O2, it is necessary to supply 110,000 g of air-saturated water, 22,000 g ofpure oxygen–saturated water, 2000 g of water containing 500 mg/L H2O2 (100% utilization), or 13 g ofair (20.9% O2)

Whether the contaminant is above or below the water table, the rate of bioreclamation in contaminated zones is effectively controlled by the rate of supply of oxygen (Wilson, Leach, Henson,and Jones, 1986) Table 5.7 compares the number of times that water in contaminated material belowthe water table, or air in material above it, must be replaced to reclaim totally subsurface materials ofvarious textures The calculations assume typical values for the volume occupied by air, water, andhydrocarbons (De Pastrovich, Baradat, Barthal, Chiarelli, and Fussel, 1979) The actual values at aspecific site will probably be different It is also assumed that the oxygen content of the water is 10 mg/L,that of the air 200 mg/L, and that the hydrocarbons are completely metabolized to carbon dioxide.After the oxygen in the air is consumed during the biological degradation of the contaminant, theremaining air should physically weather (remove volatiles by evaporation) the hydrocarbons (Wilson,Leach, Henson, and Jones, 1986) The extent of weathering depends upon the vapor pressure of thecontaminant Light hydrocarbons, such as gasoline, can be vaporized to a greater extent than they aremetabolized with oxygen The vapor pressure of gasolines varies from 100 to 1000 mm at 100°C If thevapor pressure is reduced fourfold at typical groundwater temperatures of 10°C, and benzene is typical

hydrocarbon-of the vapors, then the oxygen demand for complete metabolism hydrocarbon-of the gasoline vapors ranges from 2 to

20 times the oxygen content of air The biological and physical weathering of the hydrocarbon shouldpreferentially remove the more volatile and more water-soluble components (De Pastrovich, Baradat,Barthal, Chiarelli, and Fussel, 1979)

Substance

Application

contamination <10 mg/L COD

In situ wells Constant supply Wells subject to blow out

of oxygen possible Oxygen-enriched

peroxide

groundwater, greater oxygen concentrations can be supplied to subsurface (100 mg/L),

H2O2 provides 50 mg/L oxygen, helps to keep wells free of heavy growth

Chemical decomposes rapidly on contact with soil, and oxygen may bubble out prematurely unless properly stabilized

compounds more biodegradable

Ozone generation is expensive, toxic to microorganisms except at low concentrations, may require additional aeration

Source: U.S EPA Handbook No EPA/625/6-85/006, 1985.

Material That Originally Contained Hydrocarbons at Residual Saturation

When Drained

Air When Drained

Water When

Source: De Pastrovich, T.L et al CONCAWE Report No 3/79 The Oil Companies’ International Study Group, The Hague, The Netherlands, 1979 With permission.

Oxygen content of the soil can be improved by the presence of sand or loam (heavy clay isundesirable), avoidance of unnecessary compaction (heavy trucks, etc.), and limited loading of rapidlybiodegradable matter (Raymond, Hudson, and Jamison, 1976).

There are a multitude of chemical, photosynthetic, and electrochemical reactions that produce oxygen,either as a major or minor product (Texas Research Institute, Inc., 1982) The chemical reaction typesmost often encountered are

1 Decomposition of peroxides, superoxides (Shanley and Edwards, 1985)

(a good, ecologically sound additive, used extensively in sewage treatment; March, 1968)

Barium and strontium peroxides are used in the production of oxygenating cakes employed by fishermenfor maintaining live bait (Texas Research Institute, Inc., 1982) A typical formulation would containbarium peroxide, manganese dioxide, calcium sulfate, and dental plaster, which releases oxygen slowlywhen in contact with water However, use of materials such as these may not be advisable, because ofthe resulting heavy metal contamination of the water table Barium peroxide is definitely highlypoisonous

There is also a urea–peroxide addition compound that has been used in conjunction with phosphatesolutions to treat plants suffering from oxygen starvation in the root zone (U.S Patent 3,912,490) (TexasResearch Institute, Inc., 1982) The compound is available commercially from Western Europe It isprobably of the inclusion type, one in which H2O2 molecules are trapped within channels formed bythe crystallization of urea (March, 1968) Since the molecules are held together only by van der Waalsforces, when dissolved the solution will behave as a mixture of urea and hydrogen peroxide By weight,35% of the compound is H2O2

2 Decomposition of Peroxyacids and Salts (Austin American Statesman, 1980)

Peroxy mono- and disulfuric acids, peroxy mono- and diphosphoric acid, and peroxyborates all produceacidic solutions, but the salts may be important for consideration The exact mode of degradation ofthe salt KHSO5 (potassium monoperoxysulfate) is uncertain, but it has been used as an aid in thedegradation of atrazine (a pesticide) — presumably by virtue of its oxygen-producing ability Degra-dation probably results in the formation of KHSO4 Impure salts of peroxy monophosphoric acid(H3PO5) might prove useful

3 Thermal decomposition of oxygen-bearing salts

Generation of oxygen by this method has no particular advantage to treating underground contamination.Several alternative sources of oxygen have been suggested as a means to increase the degradative activity

in contaminated aquifers (Texas Research Institute, Inc., 1982) Oxidizing agents can be used to degradeorganic constituents in soil systems, although they may themselves be toxic to microorganisms or maycause the production of more-toxic or more-mobile oxidation products (Sims and Bass, 1984)

Two powerful oxidizing agents that have potential for in-place treatment are ozone and hydrogenperoxide

5.1.4.1 Ozone

Ozone is an oxygen molecule containing three oxygen atoms Ozone gas is a very strong oxidizing agent

that is very unstable and extremely reactive (U.S EPA, 1985a) It cannot be shipped or stored; therefore,

it must be generated on-site prior to or during application

Ozone can be employed as a pretreatment for wastes to break down refractory organics or to furnish

a polishing step after biological or other treatment processes to oxidize untreated organics (Roberts,

Koff, and Karr, 1988) It may be used to degrade recalcitrant compounds directly by creating an

oxygenated compound without chemical degradation (Texas Research Institute, Inc., 1982) Ozonation

is an oxidation process appropriate for aqueous streams that contain less than 1.0% oxidizable compounds

(Roberts, Koff, and Karr, 1988) This chemical oxidation can be used on many organic compounds that

cannot be easily broken down biologically, including chlorinated hydrocarbons, alcohols, chlorinated

aromatics, pesticides, and cyanides (Lee and Ward, 1985, 1984; Lee, Wilson, and Ward, 1987) An added

advantage of using ozone is its ability to react with PAHs (Hsu, Davies, and Masten, 1993) Ozone

venting significantly shortens remediation time and lowers residual concentrations by an order of

magnitude The rate of ozone reaction can be controlled by adjusting the pH of the medium (Texas

Research Institute, Inc., 1982) At high pH, hydroxyl free-radical reactions dominate over the more rapid

direct ozone reactions

Ozone increases the dissolved oxygen level in water for enhancing biological activity (Texas Research

Institute, Inc., 1982) The most-effective and cost-effective uses of ozone in soil system decontamination

appear to be in the treatment of contaminated water extracted from contaminated soil systems through

recovery wells, and in the stimulation of biological activity in saturated soil (Nagel et al., 1982) Ozone

treatment may be very effective for enhancing biological activity, if the organic contaminants are

relatively biodegradable However, if much of the material is relatively biorefractory, the amount of

ozone required would greatly increase the cost of the treatment

In commercially available ozone-from-air generators, ozone is produced at a concentration of 1 to

2% in air (U.S EPA, 1985a) In bioreclamation, this ozone-in-air mixture could be contacted with

pumped leachate using in-line injection and static mixing or using a bubble contact tank A dosage of

1 to 3 mg/L of ozone can be used to attain chemical oxidation However, the dosage should not be

greater than 1 mg/L of ozone per mg/L total organic carbon (TOC); higher concentrations may be

deleterious to the microorganisms At many sites, this may limit the use of ozone as a pretreatment

method to oxidize refractory organics, making them more amenable to biological oxidation

Ozone has been used to treat groundwater contaminated with oil products to reduce dissolved organic

carbon concentration (Nagel et al., 1982) Dosages of 1 g ozone/g dissolved organic carbon resulted in

residual water ozone concentration of 0.1 to 0.2 ppm The treated water was then infiltrated into the

aquifer through injection wells There was an increase in dissolved oxygen in the contaminated water

This increased microbial activity in the saturated soil zone, which stimulated microbial degradation of

the organic contaminants

Ozone was used in a petroleum-contamination incident in Karlsruhe, Germany that threatened a

drinking water supply (Atlas and Bartha, 1973c) The polluted groundwater was withdrawn, treated with

ozone, and infiltrated back into the system via three infiltration wells About 1 g of ozone per gram of

dissolved organic carbon (DOC) was added to the groundwater, with a contact time of 4 min in the

aboveground reactor (Lee and Ward, 1985; U.S EPA, 1985a) This increased the dissolved oxygen levels

to 9 mg/L, with a residual of 0.1 to 0.2 g of ozone/m3 in the treated water The dissolved oxygen reached

equilibrium at about 80% of the initial concentration injected The oxygen consumption peaked at about

40 kg/day during the initial infiltration period The microbial counts subsequently increased in the wells,

with a decrease in dissolved organic carbon and mineral oil hydrocarbons Total bacterial counts in the

groundwater increased tenfold, but bacteria potentially harmful to humans did not increase Levels of

cyanide, a contaminant identified after the treatment began, also decreased, although biodegradation was

not shown to be the cause The ozone may have also reacted with the hydrocarbon for partial destruction

of the organics The drinking water from this aquifer contained no trace of contaminants after 1 years

of ozone treatment It is conceivable that oxidizing the subsurface could result in the precipitation of

iron and manganese oxides and hydroxides If this is extensive, the delivery system and possibly even

the aquifer could become clogged

1 / 2

Saturated aliphatic compounds that do not contain easily oxidized functional groups are not readily

reactive with ozone; for example, saturated aliphatic hydrocarbons, aldehydes, and alcohols (Sims and

Bass, 1984) Reactivity of aromatic compounds with ozone is a function of the number and type of

substituents Substituents that withdraw electrons from the ring deactivate the ring toward ozone, for

example, halogen, nitro, sulfonic acid, carbonyl, and carboxyl groups Substituents that release electrons

activate the ring toward ozone, for example, alkyl, methoxyl, and hydroxyl

The following reactivity patterns with ozone are

phenol, xylene > toluene > benzene

pentachlorophenol < di-, tri-, and tetrachlorophenol

The relatively rapid decomposition rates of ozone in aqueous systems, especially in the presence of

certain chemical contaminants or other agents that catalyze its decomposition to oxygen, preclude its

effective application to subsurface waste deposits (Amdurer, Fellman, and Abdelhamid, 1985) The

half-life of ozone in groundwater is less than h (Ellis and Payne, 1984) (about 18 min; U.S EPA, 1985a)

Since the flow rates of water are likely to be in inches per hour or less, it is unlikely that effective doses

of ozone could be delivered very far for chemical oxidation However, it has been used successfully to

supply oxygen for microbial biodegradation (Rice, 1984)

Amdurer, Fellman, and Abdelhamid (1985) state that hydrogen peroxide is a weaker oxidizing agent than

ozone, but that it is considerably more stable in water It decomposes to form water and oxygen, can supply

improved oxygen levels (Lee and Ward, 1985, 1984; Lee, Wilson, and Ward, 1987), and has been used

successfully to clean up several spill sites (U.S EPA, 1985a) Advantages of hydrogen peroxide include

Greater oxygen concentrations can be delivered to the subsurface 100 mg/L H2O2 provides 50 mg/L

oxygen

Less equipment is required to oxygenate the subsurface

Hydrogen peroxide can be added in-line along with the nutrient solution Aeration wells are not

necessary

Hydrogen peroxide keeps the well free of heavy biogrowth Such growth and clogging can be a

problem in air injection systems

Hydrogen peroxide is used to degrade recalcitrant compounds and modify the mobility of some metals

(Sims and Bass, 1984) It can be used to raise oxygen levels in the soil, which can increase microbial

activity and degradation of organic contaminants (Nagel et al., 1982) Successful use of hydrogen

peroxide requires careful control of the geochemistry and hydrology of the site (Wilson, Leach, Henson,

and Jones, 1986)

Air sparging was able to maintain dissolved oxygen levels of only 1 to 2 ppm in a spill area (Nagel

et al., 1982) However, addition of microbial nutrient (a specially formulated, hydrogen peroxide-based

nutrient solution; FMC Aquifer Remediation Systems, Princeton, NJ) raised dissolved oxygen levels to

over 15 ppm This established the efficiency of hydrogen peroxide–based solutions for supplying

increased oxygen levels to enhance the bioreclamation process Hydrogen peroxide was selected as the

source of oxygen for biodegration at the Kelly Air Force Base, TX because it could provide about five

times more oxygen to the subsurface than aeration techniques (Wetzel, Davidson, Durst, and Sarno,

1986) The increase in microbial densities in stimulated underground spill sites is probably due to the

increased oxygen from the hydrogen peroxide (Wilson, Leach, Henson, and Jones, 1986b)

Hydrogen peroxide is a strong oxidant and is nonselective (Sims and Bass, 1984) It will act with

any oxidizable material present in the soil It could thus lower the concentration of natural organic

material in the soil, causing a reduced sorption capacity for some organics The effectiveness of peroxide

may be inhibited because it simultaneously increases mobility and decreases possible sorption sites,

unless this result is desired, of course

Hydrogen peroxide is effective for oxidizing cyanide, aldehydes, dialkyl sulfides, dithionates, nitrogen

compounds, phenols, and sulfur compounds (FMC Corporation, 1979) The following chemical groups

have incompatible reactions with peroxides (i.e., the reaction products are more mobile) (Sims and Bass,

1984):

1 / 2

Acid chlorides and anhydrides

Acids, mineral, nonoxidizing

Acids, mineral, oxidizing

Metals and metal compounds

Phenols and cresols

Sulfides, inorganic

Chlorinated aromatics/alicyclics

Hydrogen peroxide is more soluble in water than molecular oxygen and may provide more oxygen

at specific sites of application (Britton, 1985) The enzymatic decomposition reactions are

2H2O2→ 2H2O + O2

H2O2 + XH2→ 2H2O + Xwhere X can be NADH, glutathione, or other biological reductants

Hydrogen peroxide and ozone have been used in combination to degrade compounds that are

refrac-tory to either material individually (Nakayma et al., 1979) There is an ongoing debate as to what oxidant

is the best Some believe that hydrogen peroxide is the most-efficient way to move oxygen through a

formation Others find that air is the most cost-effective oxidizing agent On the other hand, proponents

of both hydrogen peroxide and ozone also use aeration in their bioreclamation systems

Hydrogen peroxide is reasonably inexpensive and can be produced from a coproduct process, such

as the initial conversion of glucose to gluconic acid with glucose-1-oxidase (Hou, 1982) It is

nonper-sistent and is not likely to represent a serious health hazard, if used properly (Texas Research Institute,

Inc., 1982; Britton, 1985) However, it is cytotoxic (3% is commonly used as a general antiseptic) and

may decompose (by enzymatic catalysis or nonenzymatically by in situ physicochemical processes)

before reaching its targeted spill location

This chemical can be toxic to the microorganisms it is intended to stimulate However, the growth

rate of hydrocarbon-utilizing bacteria is not necessarily inhibited by high hydrogen peroxide

concentra-tions (Texas Research Institute, Inc., 1982) Even growth enhancement is sometimes observed Whether

or not a given hydrogen peroxide concentration will be toxic to bacteria depends upon the concentration

of the organisms when the hydrogen peroxide is added Large populations are more successful at

surviving high hydrogen peroxide concentrations than are small populations

Bacteria produce hydrogen peroxide themselves from respiratory processes, and almost all aerobic

bacteria have enzymes (hydroperoxidases — catalase and peroxidase) to protect against the toxicity of

the compound (Texas Research Institute, Inc., 1983; Britton, 1985) There appears to be a critical

H2O2:organism ratio, above which the catalase-utilizing protective mechanisms of the organisms are

overwhelmed This ratio may be on the order of 2 × 1010:1 in a given volume of solution, or it may be

expressed as 1 ppm H2O2:8.9 × 105 bacteria

Catalase buildup can result in too rapid H2O2 decomposition and wasteful off-gassing of oxygen

(Spain, Milligan, Downey, and Slaughter, 1989) Britton (1985) noticed that nonviable cell material is

just as capable as the enzyme in viable cells of catalyzing decomposition of hydrogen peroxide It is

impossible to distinguish between abiotic and biotic use of the oxygen produced by this reaction (Huling,

Bledsoe, and White, 1991) Soils decompose H2O2 to the oxygen needed for in situ bioremediation by

both biotic and abiotic catalysis, and they vary greatly in peroxide decomposition activity (Lawes, 1991)

Autoclaving or treatment with acidic mercuric chloride should help estimate how much decomposition

occurs by either mode Phosphates cannot be used to estimate the extent of abiotic decomposition (Lawes,

1991)

Hydrogen peroxide has been shown to be toxic to fresh bacterial cultures at levels greater than

100 ppm, although mature cultures suffered less and could function at levels as high as 10,000 ppm.Subsequent experimentation with sand columns inoculated with gasoline and gasoline-degrading bacteriashowed that 1.0% (10,000 ppm), 0.5% (5000 ppm), and 0.25% (2500 ppm) hydrogen peroxide solutionswere toxic to the bacteria

Other studies have shown that hydrocarbon-degrading bacteria can adapt to tolerate hydrogen peroxideequivalent to 200 mg/L oxygen, a 20-fold increase in oxygen over water sparged with air (Lee and Ward,1985) In a mixed culture of gasoline degraders, the maximum concentration of H2O2 that could betolerated was 0.05% (500 ppm), although by increasing the concentration gradually, the level of tolerancecould be raised to 0.2% (2000 ppm) (Texas Research Institute, Inc., 1983; Britton, 1985) The remediation

at Grange, IN, involved adding an initial concentration of 100 ppm, and increasing it to 500 ppm overthe course of the treatment (U.S EPA, 1985a) Most applications of H2O2 have used “safe” levels,generally less than 2000 mg/L (Brown and Norris, 1994) Approximately 2 kg of hydrogen peroxide arerequired to generate 1 kg of oxygen for biodegradation of 1 kg of hydrocarbon (Norris, Dowd, andMaudlin, 1994)

There is field evidence for enhanced degradation with the use of hydrogen peroxide (Yaniga andSmith, 1984) During air sparging with 100 ppm hydrogen peroxide, the dissolved oxygen concentrations

in monitoring wells at a site contaminated by gasoline increased from 4 to 10 ppm This was accompanied

by an increase in the numbers of gasoline-utilizing organisms and a reduction in the size of the gasolineplume and a decrease from 4 to 2.5 ppm hydrocarbon However, other restoration measures wereconcurrently being employed Other authors report a greater benefit from using hydrogen peroxide insoils contaminated with JP-5 and diesel fuel than in soils contaminated with lubricating oil (Flathman,Carson, Whitehead, Khan, Barnes, and Evans, 1991) Ho, Shebl, and Watts (1995) have developed an

injection system for in situ catalyzed peroxide remediation of contaminated soil.

Hydrogen peroxide could be used in sand columns but not in batch liquid cultures (Wilson, Leach,Henson, and Jones, 1986) The liquid cultures were extremely sensitive to the compound This wasprobably due to the nature of the growth in the two environments: in sand, the organisms would grow

as a film with multicellular depth; in liquid, they would be unicellular, unprotected by adjacent cells.The rate of decomposition of hydrogen peroxide to oxygen must be controlled (Lee and Ward, 1985).Rapid decomposition of only 100 mg/L (100 ppm or 0.01%) hydrogen peroxide will exceed the solubility

of oxygen in water, resulting in bubble formation, which could lead to gas blockage and loss ofpermeability Nevertheless, it may be possible to overcome this limitation by stabilizing the hydrogenperoxide solution (Lee and Ward, 1985, 1984; Lee, Wilson, and Ward, 1987) Liberation of oxygen withresulting bubble formation should also occur in soil and groundwater with high concentrations of ferriciron (Wilson, Leach, Henson, and Jones, 1986) Addition of FeCl3 to a pumping solution would be away to form pockets of oxygen bubbles in a short time for bioreclamation of an underground gasolinespill

The hydrogen peroxide might decompose before it reaches the depths required and cause precipitation

of iron and manganese oxides and hydroxides (U.S EPA, 1985a) Much of the decomposition of hydrogenperoxide in soil and groundwater will be due to reactions with iron salts (Haber and Weiss, 1934).Nonenzymatic decomposition can occur in a variety of reactions, including those in the presence of ironsalts, known as Fenton chemistry (Fenton, 1894) The hydroxyl radical (OH–) is known as Fenton’sreagent (Ho, Shebl, and Watts, 1995) The following reactions show how different iron salts affect thedecomposition of H2O2 (Haber and Weiss, 1934)

Most decreases of hydrogen peroxide occur rapidly in the top 5.5 cm of a sand column, with onlyslight decreases thereafter, which may be due to iron stimulation (Wilson, Leach, Henson, and Jones,1986) On the one hand, the molecular oxygen produced from these reactions would help enhancegasoline biodegradation On the other hand, iron can cause the hydrogen peroxide to decompose before

it reaches the intended site The decomposition rate of peroxide is also greatly accelerated in the presence

of another heavy metal ion, Cu++ (Bambrick, 1985)

There are a number of ways to prevent the hydrogen peroxide from decomposing Standard practice

is to add enough phosphate to the recirculated water to precipitate the iron (Wilson, Leach, Henson, andJones, 1986) High concentrations of phosphates (10 mg/L) (0.01 M monobasic potassium phosphate;Britton, 1985) can stabilize peroxide for prolonged periods of time in the presence of ferric chloride,

an aggressive catalyst (U.S EPA, 1985a) The stabilizing effect of phosphate is fortuitous, since it is amajor nutrient for enhancement of underground biodegradation of gasoline How phosphates are appliedand tested are important factors in evaluating phosphates for stabilizing hydrogen peroxide (Lawes,

1991) Compounds used for in situ stabilization of peroxide in uranium mining might also improve

stabilization by phosphate (Lawes and Watts, 1981)

There are problems associated with adding high phosphate concentrations, such as precipitation (U.S.EPA, 1985a) Also, phosphates only partially protect against abiotic decomposition (Lawes, 1991) Somesuppliers add an organic inhibitor that will stabilize the peroxide at a rate appropriate to the rate ofinfiltration, so the oxygen demand of the bacteria attached to the solids is balanced by the oxygensupplied by decomposing peroxide in recirculated water (Wilson, Leach, Henson, and Jones, 1986).Dworkin Foster medium, or a similar medium containing the mineral components for growth (exceptfor a source of carbon and energy), can also stabilize hydrogen peroxide and would be a suitable solutionfor pumping the material underground without premature decomposition (Britton, 1985)

Enhancement of the microbial population has also been reportedly used to reduce levels of iron andmanganese in the groundwater (Hallberg and Martinelli, 1976) The process, known as the Vyrodexmethod, was developed in Finland and has been used in Sweden and other areas where high levels ofthe two elements are found in the groundwater Iron bacteria and manganese bacteria oxidize the solubleforms of iron and manganese to insoluble forms; the bacteria use the electrons adsorbed from theoxidation process as sources of energy Dissolved oxygen is added to the groundwater to stimulate thebacteria to first remove the iron and then later the manganese As the iron bacteria population builds upand begins to die, it supplies the organic carbon necessary for the manganese bacteria The efficiency

of the process increases with the number of aerations

Hydrogen peroxide can mobilize metals, such as lead and antimony, and, if the water is hard,magnesium and calcium phosphates can precipitate and plug the injection well or infiltration gallery(Wilson, Leach, Henson, and Jones, 1986) Heavy metal control procedures involve techniques thateffectively prevent contact between the metals and the peroxide (Bambrick, 1985) This is accomplished

by using a chelating agent and silicate The most effective of the commercially available chelating agents

is the pentasodium salt of diethylenetriaminepentaacetic acid (Na5DTPA) This is a negatively chargedcompound that can form a ringed structure that alters the reactivity of a positively charged ion Theheavy metal ion is bound by covalent bonds off the nitrogens and ionic bonds off the acetate groupsand, thus, is inhibited from entering into undesirable reactions, e.g., Na3MnDTPA The breakdown ofperoxide can be decreased substantially using the chelate Na5DTPA in combination with sodium silicateand MgSO4 The real value of DTPA, even when the metal level is low, is in stabilizing the peroxideliquor solution In the laboratory, this combination reduces the amount of peroxide decomposed to 55%after 2 h Without DTPA, 95% of the peroxide is made useless after 1 h These results have also beenverified in field tests

The pH does not strongly influence the rate of hydrogen peroxide decomposition by iron salts inaqueous media (Wilson, Leach, Henson, and Jones, 1986) The pathway of its decomposition dependsupon the valence of the iron Lawes (1991) found that an alkaline pH accelerates decomposition.Decreasing the ratio of hydrogen peroxide to soil (e.g., 1:4) also increases decomposition

The plugging of interparticular spaces in the soil resulting from the growth of biomass can lead toformation of anaerobic conditions and, in some cases, formation of toxic or explosive gases and pollutants

more toxic than the original ones (Kaufman, 1995) Prevention of biofouling during in situ bioremediation

of hydrocarbon-impacted soil or groundwater involves appropriate engineering and hydrogeologicalconsiderations prior to process initiation, as well as the judicious use of hydrogen peroxide

Although there are some compounds that will not react with hydrogen peroxide but will react with ozone

or hypochlorite, hydrogen peroxide appears to be the most feasible for in situ treatment (U.S EPA, 1985a).

Another potential oxidant is hypochlorite (Amdurer, Fellman, and Abdelhamid, 1985) It is generallyavailable as potassium, calcium, or sodium hypochlorite (bleach) and is used in the treatment of drinkingwater, municipal wastewater, and industrial waste (U.S EPA, 1985a) It reacts with organic compounds

as both a chlorinating agent and an oxidizing agent Hypochlorite additions may lead to production ofundesirable chlorinated by-products (e.g., chloroform) rather than oxidative degradation products There-

fore, the use of hypochlorite for in situ treatment of organic wastes is not recommended.

Oxygen can be supplemented with other electron acceptors, such as nitrate (Wilson, Leach, Henson, andJones, 1986) Nitrate can support the degradation of xylenes in subsurface material (Kuhn, Colberg,Schnoor, Wanner, Zehnder, and Schwarzenbach, 1985) This approach is still experimental but offersconsiderable promise because nitrate is inexpensive, is very soluble, and is nontoxic to microorganisms,although it is of human health concern Nitrate itself is a pollutant limited to 10 mg/L in drinking water(U.S EPA, 1985b) Its use may also be limited by regulations and concerns for nitrite formation andpotential for eutrophication (Brown, Mahaffey, and Norris, 1993) Section 3.2.2.1.1 describes the process

of denitrification in depth and discusses applications for bioremediation

While it has been investigated as an alternate electron acceptor for degradation of monoaromatic(except benzene) and polyaromatic compounds, nitrate does not result in degradation of aliphaticcompounds (Brown, Mahaffey, and Norris, 1993) Another study found that neither nitrate nor sulfate

as terminal electron acceptors in an anaerobic process is effective on the types of saturated hydrocarbonsfound in petroleum (Texas Research Institute, Inc., 1982)

When nitrate is the electron acceptor, 50.0 mg nitrate/L water would require 90,000 kg water/kghydrocarbon (Hinchee and Miller, 1990) With 300.0 mg nitrate/L water, 15,000 kg water/kg hydrocarbonwould be required for biodegradation

Other options for electron acceptors include carbon dioxide, sulfate (see Section 3.2.2.1.2), and iron(Brown and Norris, 1994) Iron-reducing bacteria have been shown to be able to use organic matter from

a landfill leachate as a carbon source and iron oxides in an aquifer as an electron acceptor to degradetoluene (Albrechtsen, 1994)

Cost may be the deciding factor when more than one electron acceptor would work, with hydrogenperoxide being the most expensive (Brown and Norris, 1994) There may be instances, however, whereother methods cannot be used, may require too much time, or would be ineffective Selection of theappropriate oxidizing agent depends, in part, upon the substance to be detoxified and also upon thefeasibility of delivery and environmental safety (U.S EPA, 1985a)

See Section 3.2.2 for a presentation of anaerobic biodegradation processes, biodegradable petroleumcomponents, and the microorganisms capable of degrading petroleum compounds under anaerobicconditions

The contaminated soil should remain permeable to water and air during bioremediation (NcNabb,

Johnson, and Guo, 1994) A common biological in situ remediation technique is to inject water saturated

with air oxygen, pure oxygen, or enriched with hydrogen peroxide or ozone, and sometimes nutrients,into contaminated soil (Lund, Swinianski, Gudehus, and Maier, 1991) For technical, economic, andtime reasons, the success of this approach is questionable (Lund and Gudehus, 1990)

1 Injecting Water

Oxygen must be dissolved in the interstitial water of the soil (Thibault and Elliott, 1980) Keeping thesoil moist is, in itself, a simple and low-cost method of supplying some aeration, although it providessuch a low amount of oxygen, it is not very effective

2 Colloidal Gas Aphrons

A newly developed method that holds great promise for introducing oxygen to the subsurface ismicrodispersion of air in water using colloidal gas aphrons (CGA), which creates bubbles 25 to 50 µm

in diameter (U.S EPA, 1985a) With selected surfactants, dispersions of CGAs can be generatedcontaining 65% air by volume A surfactant concentration of 1000 to 5000 ppm is needed to generatethe microbubbles (Lange, Bouillard, and Michelsen, 1995) Foam, in the form of microbubbles, consists

of 60 to 70% dispersion of 55-µm microbubbles in water (CGA) Foam flows into less accessible spillareas, without channeling or poor sweep

3 Injecting Air

Air is much less viscous than water (1.8 × 102 and 1.0 × 104 µP, respectively) (Wilson, Leach, Henson,and Jones, 1986) Air also has a 20-fold greater oxygen content on a volume basis If the air- and water-filled porosity are about the same, and the pressure gradients are the same, then air should be about

1000 times more effective than water Air should be particularly effective for oxygen supply to inated regions high in the unsaturated zone

contam-Air can be injected by use of pumps, propellers, stirrers, spargers, sprayers, and cascades (Texas ResearchInstitute, Inc., 1982) A blower can be used to provide the flow rate and pressure for aeration, such as 5 psipressure in a 10-ft aeration well, with an airflow of 5 ft3/min (Texas Research Institute, Inc., 1982).Air can also be added to extracted groundwater before reinjection, or it can be injected directly into

an aquifer (U.S EPA, 1985a) The first method, in-line aeration, involves adding air into the pipelineand mixing it with a static mixer to provide a maximum of 10 mg/L oxygen This concentration willdegrade about 5 mg/L hydrocarbons and would, therefore, provide an inadequate oxygen supply Apressurized line can increase oxygen concentrations, as can the use of pure oxygen

4 Oxygenation Systems

Oxygenation systems, either in-line or in situ, can also be installed to supply oxygen (U.S EPA, 1985a).

These can achieve higher oxygen solubilities and more-efficient oxygen transfer to the microorganismsthan conventional aeration Solubilities of oxygen in various liquids are four to five times higher underpure oxygen systems than with conventional aeration In-line injection of pure oxygen can impart 40 to

50 ppm of dissolved oxygen to water (Brown, Norris, and Raymond, 1984), which will provide sufficientdissolved oxygen to degrade 20 to 30 mg/L of organic material, assuming 50% cell conversion Thisoxygen will not be consumed immediately, as is the oxygen from aeration Pure oxygen is expensive touse and the oxygen is likely to bubble out of solution (degas) before the microbes can utilize it.Pure oxygen can be injected by use of pumps, propellers, stirrers, spargers, sprayers, and cascades(Texas Research Institute, Inc., 1982)

5 Injection of Liquified Gases

The injection of liquid oxygen or liquid air into the soil could utilize existing technology (Texas ResearchInstitute, Inc., 1982) Intermittent injection of liquid oxygen would produce a high concentration ofoxygen, which would slowly diffuse into the surrounding strata Since oxygen is ten times more soluble

in hydrocarbons than it is in water, the hydrocarbon phase could actually act as an oxygen reservoir toreplace the oxygen being consumed in the aqueous phase (Faust and Hunter, 1971) Repeated injectionswould create a flow through the system, preventing buildup of carbon dioxide Another technique wouldhave to be used to add additional nutrients This method would be best in an area where the soil containedabundant nutrients

Liquid oxygen was trucked to a site contaminated with PAHs (Gupta, Djafari, and Zhang, 1995) Aspacing of about 12 m was used between infiltration system laterals and for extension galleries Withthe proposed infiltration trench and overlying collection gallery design for the waste matrix, fluid seepagevelocities exceeded 2 m/day, with a minimum of 4 mg/L dissolved oxygen concentration conducive forbioremediation

A technique for soil venting was used in Mont Belvieu, TX to flush leaked propane and ethane out

of the ground (Austin American Statesman, 1980) Liquid nitrogen was pumped underground, and the

large volumes of nitrogen gas generated swept the gases through the soil Possibly, liquid oxygen or aircould be utilized in this fashion to supply oxygen to the soil strata It is not known what effect a strayspark or flame might have on a system such as this The potential for an underground fire exists

6 Injection of Oxygen-Releasing Compounds (with Nutrients)

The best material for implementing this approach is hydrogen peroxide (Texas Research Institute, Inc.,1982) Injection should ideally be made over the entire contaminated area, both into the water table and

at points just above the water table into the gasoline-bearing soil A large amount of the residual gasoline

would be consumed by the bacteria in the soil The organisms may produce emulsifiers that would helpmobilize the gasoline into the water table where it could be collected at the producing wells Arecirculating system might be set up to treat the produced water by cleaning, fertilization, oxygenation,and reinjection into the water table Oxygen is best provided through recirculated groundwater usinghydrogen peroxide, if it has to be delivered to fractured bedrock, highly stratified aquifers, or where thesaturated interval is 1 m or less (Brown, Mahaffey, and Norris, 1993).

A variation would be to use a physical oxygenation technique on the injection water instead of achemical additive See Section 5.1.4.2 for a discussion of the use of hydrogen peroxide for bioremediation

7 Well Points

Site geology in most cases will determine the methods of aeration to be used in a given situation(Raymond, Jamison, and Hudson, 1976) For example, in a fractured dolomite and clay formation, lack

of homogeneity makes well injection and distribution of oxygen difficult

A well point injection system can be used to supply oxygen and nutrients to the subsurface Air can

be sparged into wells using diffusers Diffusers attached to paint sprayer-type compressors can injectabout 2.5 cfm air into a series of wells to enhance degradation (Raymond, Jamison, and Hudson, 1976;U.S EPA, 1985a) The diffusers are positioned 5 ft from the bottom of the well and below the watertable Aeration through well points has been successfully used for saturated soils, but it is uncertainwhether or not the technique would also work for unsaturated soils

The solubility of oxygen is very low, approximately 8 mg/L at groundwater temperatures (Wilsonand Rees, 1985) Diffusers that sparge compressed air into the groundwater cannot exceed the solubility

of oxygen in water (Lee and Ward, 1986) Oxygen levels can be increased by sparging the injectionwells with oxygen instead of air (Wilson, Leach, Henson, and Jones, 1986) This raises the oxygen levelsabout fivefold

8 Shallow Injection Sites

Since a well or injection site is expensive to drill, relatively shallow soil injection sites that are just deepenough to get oxygen and nutrients past the plant growth zone may be adequate (Texas Research Institute,Inc., 1982)

9 Plowing/Tilling

Even though spills may contaminate just the upper layer of soil, an oxygen limitation can retarddegradation (Thibault and Elliott, 1980) Active microflora have been observed in the top 15 cm of soil,and tilling is suggested as an effective means of promoting aeration (Raymond, Hudson, and Jamison,1976) This provides aeration by turning over the soil layers with plows and exposing the soil to theatmosphere to a depth of 6 to 24 in (Ju, Devinny, and Paspalof, 1993) Traditional machines for soilplowing are disk harrows and rototillers Soils deeper than about 2 ft can be aerated by using constructionequipment, such as a backhoe Tilling equipment can aerate surface soils and mix wastes or reagentsinto the soil (Sims and Bass, 1984) Rototilling equipment promotes the aeration and mixing processmore effectively than disks or bulldozers (Raymond, Hudson, and Jamison, 1976)

Plowing may be valuable for providing aeration to greater depths in the active layer of the soil, butthis is only the case when soils are very active, and it is unlikely to maintain soils in aerobic conditionswhen this is the case (Devinny and Islander, 1989) Although plowing will ensure adequate mixing ofwater and fertilizer, plowed soil still remains highly heterogeneous (Ju, Devinny, and Paspalof, 1993).Plowing will break up large clods but may not affect small or hard aggregates, which prevent penetration

of oxygen and protect entrapped petroleum hydrocarbons from degradation Total petroleum hydrocarbonconcentrations are higher in soils containing aggregates Tillage is effective to only a depth of about

15 cm and destroys soil structure of wet soil, forming large clods and lowering the rate of bioremediation(NcNabb, Johnson, and Guo, 1994)

For a particular field experiment, rent for a disk harrow was $150/h (Ju, Devinny, and Paspalof, 1993).This equipment could plow about 20 ha/h but required two or three passes Assuming two passes(10 ha/h), rental costs would be $15/ha for each treatment With labor cost of $30/h, total treatment costswould be $18/ha If the disked site were treated once a week for 6 weeks, the total cost would be $108/ha

10 Pulverization

The time required for bioremediation can be reduced by substituting pulverization for the traditionalplowing to aerate the contaminated soil (Ju, Devinny, and Paspalof, 1993) Experimentation determined

that oil and grease elimination rates were higher for smaller particles, consistent with the greateravailability of substrate in these particles The numbers of microorganisms may not increase on pulverizedparticles, but the biodegradation of each organism will be more rapid because of the more easilydegradable substrate exposed to them It was unclear whether pulverization would affect total volatilesrelease A 25% reduction in treatment time was observed in a field test of the pulverized system.Costs of pulverization are modest (Ju, Devinny, and Paspalof, 1993) In a field test comparing pulver-ization with plowing, the pulverizer rented for about $260/h and could treat about 4 ha/h Rental was about

$65/ha, or $68/ha with labor The pulverized site needed only four treatments, for a total of $272/ha

11 Combined Air–Water Flushing

Oxygen can be better dispersed through the subsoil by creating nearly horizontal airflow through anetwork of positive (injection) and negative (suction) lances in the soil (Lund, Swinianski, Gudehus,and Maier, 1991) In addition to this, water percolating vertically downward from a sprinkling system

on the surface detaches and solubilizes the hydrocarbons and provides moisture to the organisms for thebiodegradation Nutrients can also be dispersed with the water Both air and water flow through the soilpores During biodegradation, depleted oxygen in the water is replaced by diffusion from the air, andcontaminants are removed in the air and water streams (see Section 2.2.2.7)

12 Hydraulic/Pneumatic Fracturing

Hydraulic fractures can be created in the subsurface and utilized for more even distribution of materialsthrough the soil to enhance biodegradation (Davis-Hoover, Murdoch, Vesper, Pahren, Sprockel, Chang,Hussain, and Ritschel, 1991)

Fluid is injected into a well until the pressure nucleates a fracture A proppant (a material that propsthe fracture) such as sand can be released into the fracture, at the time of formation Granules of slow-release nutrients or oxygen-generating compounds in a viscous gel could then be pumped through thepermeable sand conduits Multiple layers of horizontal or vertical fractures can be created Although anencapsulated solid peroxide proved to be toxic when dispersed with this approach, the method is stillpromising as a delivery system for appropriate materials A test employing pneumatic fracturing at TinkerAir Force Base, Oklahoma City, OK successfully increased permeability of the unsaturated zone (Ander-son, Peyton, Liskowitz, Fitzgerald, and Schuring, 1995) Postfracture airflows were 500 to 1700% higherafter the treatment See Section 2.2.2.5

13 In Situ Electrobioreclamation/Electro-osmosis

Since contaminated soils may have a low permeability, it is difficult to deliver nutrients, molecularoxygen, other electron acceptors, or electron donors through the vadose zone (Lageman, Pool, vanVulpen, and Norris, 1995) Movement of these materials can be accelerated through the use of electro-kinetic transport

The process of electrobioreclamation has been developed for low-permeability, unsaturated soils(Lindgren and Brady, 1995; Lageman, Pool, van Vulpen, and Norris, 1995) Alternating anode andcathode arrays induce an electrical current into the soil matrix and cause soil water to flow toward thecathode (electro-osmosis) The ionic pollutants dissolved in the soil–water solution migrate and collectaround the oppositely charged electrode, where they are captured and removed Production of electricityheats the soil, which increases hydraulic permeability and the solubility of organic constituents (Lageman,Pool, van Vulpen, and Norris, 1995)

The electrodes can be installed at any depth for in situ or ex situ electroreclamation (Lageman, Pool,

van Vulpen, and Norris, 1995) Soil moisture can be controlled for optimum results Because unsaturatedconditions (typically <50% saturation) are maintained, there is a residual void space that could be utilizedfor dispersing gases (Hazen, Lombard, Looney, Enzien, Doughtery, Fliermans, Wear, and Eddy-Dilek,1994) An enclosed system would permit control of temperature, moisture, pH, oxygen, nutrients, addition

of surfactants, supplementation of highly efficient contaminant-degrading microorganisms, and toring of reactions and conditions, resulting in a more rapid and greater extent of biological degradation.This method is less dependent on favorable weather conditions, and seasonal fluctuations would not be

moni-a limiting fmoni-actor See Section 2.2.2.8 for a full description of this technology

14 Lowering the Water Table

There is another treatment approach worth considering If preliminary remediation has removed anyhydrocarbons floating on the water table, and if the geology is favorable, then it might be possible to

lower the water table to bring the entire contaminated soil into the unsaturated zone where it can bepermeated by air (Wilson, Leach, Henson, and Jones, 1986) Dewatering the soil to eliminate saturatedconditions and increase air/contaminant contact might improve biodegradation in these areas (Hinchee,Ong, Miller, Vogel, and Downey, 1992).

Air sparging alone could maintain dissolved oxygen levels of only 1 to 2 ppm in a spill area (Nagel

et al., 1982) However, when it was combined with addition of a hydrogen peroxide–based nutrientsolution (FMC Aquifer Remediation Systems, Princeton, NJ) dissolved oxygen levels rose to over

15 ppm

16 Venting (Bioventing)

Bioventing is the process of aerating subsurface soils to stimulate in situ bioremediation, while

simul-taneously removing volatile compounds (Hinchee, Ong, Miller, Vogel, and Downey, 1992) This processuses soil vapor extraction (SVE) systems to transport oxygen to the subsurface, where microorganismsare stimulated to metabolize fuel components aerobically (Dupont, 1993) Bioventing systems aredesigned to optimize oxygen transfer and utilization and are operated at much lower flow rate and withconfigurations much different than those of conventional SVE systems Lower flow rates allow longervapor retention times in the soil This promotes greater biodegradation (Miller, Vogel, and Hinchee,1991) and reduces the amount of discharged vent gas requiring treatment (Dupont, Doucette, andHinchee, 1991) Airflow rates can be adjusted to maintain oxygen levels between 2 and 4% for thispurpose (Miller, Vogel, and Hinchee, 1991) An airflow rate of 0.5 air void volumes per day was found

to be optimal at one test site See Sections 2.2.1.11 and 2.2.2.2 for a full description of the processes

of soil venting and bioventing

Variations of the soil venting technique are being investigated (Downey, Frishmuth, Archabal, Pluhar,Blystone, and Miller, 1995) One alternative involves low rates of pulsed air injection, a period of high-rate SVE, and off-gas treatment followed by long-term air injection An innovative remediation approach

that combines regenerative resin ex situ vapor treatment with in situ bioventing to reduce overall costs

of site remediation has been developed (Downey, Pluhar, Dudus, Blystone, Miller, Lane, and Taffinder,1994) Cyclic, or surge, pumping vents air to the organisms as needed (Dupont, 1993) The retainedvapors have a longer exposure to microbial degradation in the soil Bioventing is being combined withbioslurping, a vacuum-enhanced, free-product recovery system for light nonaqueous-phase liquids(LNAPLs) (Kittel, Hinchee, Hoeppel, and Miller, 1994) In this process, LNAPLs are recovered whilethe vadose zone is biovented (see Section 3.2.1.6)

17 Forced Aeration with Biopiles

Biodegradation of organic contaminants by indigenous microorganisms can be stimulated by ex situ

forced aeration of soil piles (Battaglia and Morgan, 1994) Slotted pipes extend through the pile and areattached to a vacuum blower, which draws air through the soil, aerating the microorganisms Straw,sawdust, or manure can improve soil permeability Without forced air, the piles would require a longertreatment time, as well as more space, although this would eliminate the need for treating off-gases(Benazon, Belanger, Scheurlen, and Lesky, 1995) This technology is fully described in Section 2.1.2.1.4

18 Lagoons and Waste Stabilization Ponds

Lagoons will require aeration and additional nutrients for effective petroleum biodegradation (McLean,1971) Aerating accelerates biodegradation (Johnson, 1978), but it may also be necessary to buffer thewater and include algae to provide continuous oxygen

19 Sludge Systems

More-rapid breakdown of chemicals can be accomplished by utilizing pure oxygen or oxygen-enrichedair instead of air in activated sludge systems (Roberts, Koff, and Karr, 1988) Extended aeration involveslonger detention times than conventional activated sludge Addition of pure oxygen raised performance

by 10% (Lopatowska, 1984)

20 Bioreactors

Bioreactors allow tighter control of oxygen levels during treatment of contaminated soil than can be

achieved in situ The range of 50 to 80% of the maximum water capacity provides high oxygen

consumption in bioreactors, but may vary with the particular soil type (Stegmann, Lotter, and klage, 1991) There are many ways that air or oxygen can be added to the system, depending upon thetype of reactor used For instance, air can be added intermittently to soil-slurry reactors or by bubblelessoxygenation with a membrane gas-transfer system to prevent foaming (Stormo and Deobald, 1995).Check the individual processes in Section 2 for more specific information

1 Bio XL/Restore

The Aquifer Remediation System Bio XL process employs stabilized solutions of hydrogen peroxide(tradename Restore) to increase the amount of oxygen in the soil by more than 25 times, in comparisonwith air sparging (Chowdhury, Parkinson, and Rhein, 1986) Another of its products (Restore MicrobialNutrient) prevents precipitation of chemical nutrients Bioreclamation can now be used in low-perme-ability formations, where the pumping rate from recovery wells is as low as 5 gal/min

2 Enhanced Natural Degradation

Groundwater Technology has a similar in situ process, called END (Enhanced Natural Degradation) It

is planning to introduce a new system that could cut the amount of hydrogen peroxide consumption by

75 to 90% by modifying the oxygen delivery system into a closed loop

3 Tilling

There can be severe oxygen limitation to degradation within inches of the surface of soil (Zitrides, 1983).Polybac Corporation employs tilling of the soil to provide additional oxygen, as well as to better mix amicrobial inoculum with the contaminant Otherwise, the organisms will adhere to the top layers of soiland percolate only slowly to greater depths

Use of oxygen in the Mixflo system reduces the off-gas volume by over 99%, compared with othertechnologies using air (Bergman, Greene, and Davis, 1994) By reducing off-gas volumes, this systemminimizes the problem of aqueous, oily organic wastes foaming during bioremediation and minimalemission being released to the atmosphere

The low-volume airflow of bioventing is combined with a closed-loop concept to regulate soil moisture,nutrients, and oxygen (Burke and Rhodes, 1995) Vapors are extracted via wells and treated on-site.Oxygen, nutrients, heat, and moisture are added to the vapor stream, which is then reinjected into thecontaminated soil above the water table, with no off-gases to treat The system is called BioPurgeSM,when the vapor is injected above groundwater, and BioSpargeSM, when it is injected below groundwaterlevel It is primarily for use in permeable soils See Section 2.2.2.4 for a full description of the process

7 Deep Soil Fracture Bioinjection™

Deep Soil Fracture Bioinjection™ uses pressurized subsurface injection, in an overlapping grid pattern,

of a slurry containing controlled-release nutrients, oxygen, and microbes (Burke and Rhodes, 1995)

Injected materials permeate all types of soil through desiccation cracks, fissures, and sand lenses toensure excellent transmittance, even in silt and clay, to depths of 40 ft It is an effective and economicaltechnique The process is further described in Section 2.2.2.6.

8 Vacuum Heap Biostimulation System

The Ebiox bioremediation system employs natural, contaminant-adapted microorganisms for large-scaletreatment of excavated soil (Eiermann and Menke, 1993; Eiermann and Bolliger, 1995a) Contaminatedsoil is excavated and piled in several layers on a sealed biobed, with perforated plastic piping runningbetween the layers and connected to a vacuum blower system A vacuum draws outside air through thesoil by bioventing to provide oxygen to the microorganisms The vacuum heap is covered by a blackplastic liner, and vapors are treated by biofilter The leachate is collected, purified, enriched with oxygenand nutrients, and resprayed over the heap (see Section 2.1.2.1.5)

9 Detoxifier™

The Detoxifier™, by Toxic Treatments (U.S.A.), Inc., San Mateo, CA, is an adaptation of drillingtechnology that can be used to deliver treatment agents to the soil to a possible depth of 60 ft (25 ft hasbeen successfully demonstrated) (Ghassemi, 1988) A process tower contains two cutter/mixer bitsconnected to separate, hollow Kelly bars, through which materials can be added to the soil in dry, liquid,slurry, or vapor form, with thorough mixing and homogenization of a vertical column of soil Air, oxygen,

or hydrogen peroxide could be easily added to the soil A metal shroud captures off-gases, which areprocessed in a treatment train and recycled to the soil On-line monitoring permits good process controland adjustment in treatment conditions The system establishes a closed-loop operation, and it is

completely mobile The process can be operated in situ, using a treatment grid pattern, or ex situ in

reactors Sections 2.2.1.13 and 8.4 provide a full description of the process

Both the compounds to be degraded and the specific microorganisms should be considered, whenassessing whether anaerobic conditions are to be employed for bioremediation Also, the size of thearea, monitoring requirements, personnel costs, and remoteness of the site must be included in theevaluation (Norris, 1995)

Anaerobic degradation can occur immediately, or it can require an acclimation period of up to

18 months (Sahm, Brunner, and Schoberth, 1986; Alexander, 1994) When oxygen is consumed fasterthan it can be replaced (e.g., if oxygen is depleted during bioremediation), there will be a shift in thecomposition of the microflora (JRB Associates, Inc., 1984) The predominant organisms will then befacultative anaerobic bacteria, which use alternative electron acceptors (e.g., nitrate) and are able toadapt from aerobic to anaerobic conditions (U.S EPA, 1985a) There will also be strict anaerobes thatcannot tolerate oxygen and would be inhibited or killed by oxygen treatment or the presence of oxidizedmaterials (Fulghum, 1983) Both fastidious anaerobes, the organisms capable of living under anoxic,but not necessarily reduced, environmental conditions, and facultative microbes are important in anaer-obic degradation (Kobayashi and Rittmann, 1982) Many soil bacteria will grow under anaerobic con-ditions, but most fungi and actinomycetes will not (Parr, Sikora, and Burge, 1983)

Active nitrate-respiring microorganisms are found in a variety of anoxic environments, includingsoils, lakes, rivers, and oceans (Berry, Francis, and Bollag, 1987) When changing the soil from anaerobic to an anaerobic status, facultative microorganisms, such as the denitrifiers, can adapt to nitraterespiratory metabolism (Gottschalk, 1979), utilizing many of the same compounds as they did underaerobiosis (Casella and Payne, 1996) Bacterial denitrification is the most effective type of biotransfor-mation in polluted, anaerobic environments, especially if nitrate or other N oxides are naturally available

or added Nitrate is more soluble and often less expensive than oxygen, although it is itself a pollutant,limited to 10 ppm in drinking water When soil contaminated with JP-4 jet fuel was enhanced withnitrate, the total number of denitrifiers increased by an order of magnitude (Thomas, Gordy, Bruce,Ward, Hutchins, and Sinclair, 1995)

Denitrification can begin when the concentration of oxygen drops below 10 µmol/L (Roberston andKuenen, 1984) Oxygen levels can be decreased by compacting the soil or by saturating the soil withwater, which restricts the diffusion of gases (Sims and Bass, 1984) Soil structure and texture primarilydetermine the size of soil pores, and hence the water content at which gas diffusion is limited and therate at which anaerobiosis occurs (JRB Associates, Inc., 1984) Reducing pore size and restricting aeration

increases anaerobic microsite frequency in the soil Compaction helps draw moisture to the soil surface.This lessens the problems of any leaching that may occur, if anaerobiosis is achieved by addition ofwater Volatilization may also be suppressed by surface soil compaction Water may still have to beadded to reach the required degree of anaerobiosis; however, it would be less than for an uncompactedsoil, also minimizing the leaching potential Diking is a common agricultural practice that may beapplicable to decreasing the soil oxygen content (Arthur D Little, 1976) This would establish andmaintain anaerobic conditions as long as the land is kept under water If the soil is heavily overloadedwith contaminants, it may become anaerobic and remain so for a long time.

If the soil contains montmorillonite, addition of moisture or water would cause the soil to swell andblock any further water movement (Hornick, 1983) Runoff or flooding could result, inducing anaerobicconditions If coarse soils of sand and gravel, with their large interconnecting pores, are excessivelydrained, nutrients in added material will not be sufficiently adsorbed on the soil The groundwater could

be contaminated, if there is no restrictive layer between the coarse layer and the water table

Another possible method of rendering the site anaerobic would be to add excessive amounts of easilybiodegradable organics so the oxygen will be depleted (U.S EPA, 1985a) An example of this is addition

of starchy potato-processing waste materials, which promote growth of the aerobic organisms until theavailable oxygen is depleted and anaerobiosis established (Stevens, Crawford, and Crawford, 1991;Kaake, Roberts, Stevens, Crawford, and Crawford, 1992) Contaminants may be immobilized in the soil

to allow more time for degradation, by controlling soil moisture content If anaerobic decomposition isdesired, anaerobiosis must be achieved by a means other than flooding, such as soil compaction ororganic matter addition Control of soil moisture may be achieved through irrigation, drainage, or acombination of methods

There can be economic advantages of using anaerobic degradative processes, since plugging andintensive management associated with hydrogen peroxide addition would be avoided However,exopolysaccharide (EPS) can lead to subsurface plugging under denitrifying conditions

The type of cometabolite (carbon source) used to stimulate biomass production under denitrifyingconditions will influence the amount of EPS production, which could lead to subsurface plugging (Han-neman, Johnstone, Yonge, Petersen, Peyton, and Skeen, 1995) Both total biomass production and growthefficiency are inversely correlated with EPS production under aerobic or anaerobic conditions Theoxidation state of the carbon source can be used to estimate the potential for EPS production Substratesmore reduced than glucose yield more energy that can be used in synthesis of polysaccharide; substratesmore oxidized than glucose allow energy to be used for cell synthesis rather than large quantities of EPS.See Section 5.1.7 for a discussion of oxidation-reduction potential and how it relates to creating ananaerobic environment

Some compounds require aerobic conditions for biodegradation, while others need anaerobic conditions.Some can be degraded under either condition, while others are not transformed at all Compounds thatappear to be resistant to anaerobic degradation include anthracene, naphthalene, benzene, aniline, 4-tolu-idine, 1- and 2-naphthol, pyridine, and alkanes (Alexander, 1994) While nitrate may serve as analternative electron acceptor for degradation of monoaromatic (except benzene) and polyaromatic com-pounds, it does not promote the degradation of aliphatic compounds (Brown, Mahaffey, and Norris,1993) Neither nitrate nor sulfate, as terminal electron acceptors, was effective on the types of saturatedhydrocarbons found in petroleum (Texas Research Institute, Inc., 1982) Alkylbenzenes in groundwatercan be transformed both aerobically and anaerobically (Wilson, Bledsoe, Armstrong, and Sammons,1986) Anaerobic degradation may be a useful adjunct to the aerobic degradation in heavily contaminatedareas of soil where the oxygen supply has been depleted

In general, hydrocarbons are subject to both aerobic and anaerobic oxidation (Dietz, 1980) The firststage of biodegradation of insoluble hydrocarbons is predominantly aerobic, while the organic carboncontent is then reduced by anaerobic action Interactions between different microbial populations can

be essential for the degradation of complex compounds (Hornick, Fisher, and Paolini, 1983) Sometimes,products from the anaerobic process require a final aerobic treatment (Alexander, 1994)

Many compounds can be transformed under anaerobic or alternating anaerobic/aerobic conditions(Wilson and Wilson, 1985), but not readily under strict aerobic conditions Sequencing anaerobic andaerobic biotreatment steps can be a viable alternative treatment approach (Field, Stams, Kato, and Schraa,

1995) Anaerobic and aerobic organisms could be combined in a single treatment step, by immobilizingthe microbes in biofilms or soil aggregates, within which anaerobic microniches can form.

The redox conditions can be controlled to achieve conditions under which specific compounds can

be degraded, dehalogenated, or particular organisms or enzyme systems can be selected (Wilson andWilson, 1985) Alteration of aerobic/anaerobic conditions by adjusting Eh through flooding or cultivationcan, therefore, be a useful tool for engineering management to maximize detoxification and degradation

of some compounds (Guenther, 1975)

Mixed oxygen/nitrate electron acceptor conditions showed that cultures could degrade benzene,toluene, ethylbenzene, and naphthalene (2 mg/L) under conditions of 2 mg/L oxygen and high levels ofnitrate (>150 mg/L NO3) (Wilson, Durant, and Bouwer, 1995) Biodegradation was inhibited at high(8.6 mg/L) and low (<2 mg/L) oxygen levels, except for toluene, which was degraded under anaerobicdenitrification conditions When Miller and Hutchins (1995) combined nitrate, 27 to 28 mg(N)/L, withlow levels of oxygen, 0.8 to 1.2 mg O2/L, they observed complete removal of toluene and partial removal

of ethylbenzene, m-xylene, and o-xylene with nitrate as the only electron acceptor The soil type, however,

caused variable results

The concentration of the contaminants affects the degradation (Wilson, Leach, Henson, and Jones,1986) The critical concentration of dissolved oxygen for promotion of aerobic activity ranges from0.2 to 2.0 mg/L and is usually 0.5 mg/L (U.S EPA, 1985a) The solubility of benzene (1780 mg/L) ismuch greater than the capacity for its aerobic degradation in groundwater (Wilson, Leach, Henson, andJones, 1986) However, anaerobic processes will often take over in these situations

Most filamentous fungi are aerobic, and yeasts are often facultatively anaerobic Some of the purplenonsulfur organisms can grow microaerobically and anaerobically as phototrophs, yet live as heterotrophsaerobically in the dark, by respiratory metabolism of organic compounds (Stanier, Doudoroff, and

Adelberg, 1970; Pfennig, 1978a) Cyanobacteria and Chlorella tolerate low oxygen levels (Kobayashi

and Rittmann, 1982) These authors developed a protocol for separating and treating refractory pounds by using the bioaccumulation capability of phototrophs (Sections 2.1.2.2.1.4 and 5.2.1.5)

com-Aerobic/anaerobic degradation is used to treat easily degradable constituents Refractory compounds areremoved by sorption to microorganisms, concentrated, and disposed of by other means, such as burial

or incineration Relatively recalcitrant compounds can be degraded if they are first sorbed to thetic organisms The products of these reactions could then be further degraded by other microorganisms.Aerobic or anaerobic conditions can be selected to encourage degradation by the most favorableprocess for the compounds of concern A combination of the two may sometimes allow the most completebiodegradation to occur A variety of methods is listed for modifying soil oxygen content to achieveaerobic (Sections 5.1.4.5 and 5.1.4.6) and anaerobic (Section 5.1.4.7) conditions

photosyn-BARR (bioanaerobic reduction and reoxidation) is a remedial technique for in situ degradation of

organics in soil and groundwater employing broad genera, conditioned microorganisms (Dieterich, 1995).The process provides substrate and controlled aeration to create strong shifts in redox to destabilize

contaminants and improve in situ biodegradation by means of direct metabolism, cometabolism, and

surface catalysis

If a surfactant is required, Bacillus licheniformis JF-2 synthesizes a surfactin-like lipopeptide that is

the most effective biosurfactant known (Lin, Carswell, Georgiou, and Sharma, 1994) This peptide isproduced under both aerobic and anaerobic conditions, but can be optimized by growing the cells under

O2-limiting conditions and a low dilution rate of 0.12/h

Microbial degradation of hazardous compounds requires the presence of nitrogen, phosphorus, andpotassium, in addition to smaller levels of zinc, calcium, manganese, magnesium, iron, sodium, andsulfur for optimum biological growth (Arora, Cantor, and Nemeth, 1982) Nitrogen and phosphate arethe nutrients most frequently present in limiting concentrations in soils (U.S EPA, 1985a) The nutrientsrequired by microorganisms are presented in Table 5.8 (Alexander, 1977)

Feeding nutrient solutions containing inorganic nutrients, such as nitrogen, phosphorus, and sulfur,

to natural soil bacteria often enhances the ability of the microorganisms to degrade organic moleculesinto carbon dioxide and water (Stotzky and Norman, 1961a; 1961b) Even adding cane sugar molasses

to Omani crude oil increased microbial respiration and n-alkane biodegradation (Al-Hadhrami,

Lappin-Scott, and Fisher, 1996) At one site, decomposition of oil in the soil was shown to proceed at a rate of

0.5 lb/ft3/month without a nutrient source, and at 1.0 lb/ft3/month after the addition of fertilizer cannon, 1972).

(Kin-Normally, the aliphatics are more easily degraded than aromatics (Perry and Cerniglia, 1973) Withoutadded nutrients, aromatic hydrocarbons are noted to be more readily attacked than saturated aliphatichydrocarbons by the microbes (Atlas, 1981) Addition of nitrogen or phosphorus stimulates degradation

of saturated hydrocarbons more than that of aromatic hydrocarbons Aromatics might also exhibit morerapid degradation than aliphatics, if the soil contains more organisms with a preference for the former(Zhou and Crawford, 1995) However, addition of nutrients increases total biomass available for degra-dation of all compounds and reduces the difference noticed

Some of these nutrients may be present in contaminating wastes, but may not be readily available or

in the amount required (Sims and Bass, 1984) Their supplementation may be necessary Various authorsprovide ratios for these nutrients There is some disagreement on the exact ratios, but they are not toodissimilar and a range will be provided here Three of the major nutrients, nitrogen, phosphorus, andpotassium, can be supplied with common inorganic fertilizers (JRB Associates, Inc., 1984) The carbon,nitrogen, and phosphorus content of bacterial cells is generally in the ratio of 100 parts carbon to 15 partsnitrogen to 3 parts phosphorus (Zitrides, 1983) Theoretically, 150 mg of nitrogen and 30 mg of phos-phorus are required to convert 1 g of hydrocarbon to cellular material (Rosenberg, Legmann, Kushmaro,Taube, Adler, and Ron, 1992) By knowing how much of the carbon in a spilled substance ends up asbacterial cells, it is possible to calculate the amount of nitrogen and phosphorus necessary to equal thisratio for optimum bacterial growth (Thibault and Elliott, 1980)

Measurement of soil organic carbon, organic nitrogen, and organic phosphorus allows the nation of its carbon-to-nitrogen-to-phosphorus (C:N:P) ratio and an evaluation of nutrient availability(Sims and Bass, 1984) If the ratio of organic C:N:P is wider than about 300:15:1, and available(extractable) inorganic forms of nitrogen and phosphorus do not narrow the ratio to within these limits,supplemental nitrogen and/or phosphorus should be added, such as by addition of commercial fertilizers(Kowalenko, 1978) One such product is POLYBAC® N Biodegradable Nutrients, which contains theproper balance of nitrogen and phosphorus required in a form readily available for microbial uptake(Thibault and Elliott, 1979) Sufficient nitrogen and phosphorus should be applied to ensure that thesenutrients do not limit microbial activity (Alexander, 1981)

determi-The C:N ratio might be the primary factor in determining the nutrient effect (Zhou and Crawford,1995) The quantity of nitrogen and phosphorus required to convert 100% of the petroleum carbon tobiomass may be calculated from the carbon-to-nitrogen (C:N) and carbon-to-phosphorus (C:P) ratiosfound in cellular material (Dibble and Bartha, 1979a) Accepted values for a mixed microbial population

in the soil are C:N, 10:1 (Waksman, 1924); and C:P, 100:1 (Thompson, Black, and Zoellner, 1954) Inreality, a complete assimilation of petroleum carbon into biomass is not achievable under naturalconditions Some of the petroleum compounds are recalcitrant or are metabolized slowly over longperiods From petroleum compounds that are readily metabolized, some carbon will be mineralized to

1 Energy source Organic compounds

Inorganic compounds Sunlight

a Amino acids Alanine, aspartic acid, glutamic acid, etc.

b Vitamins Thiamine, biotin, pyridoxine, riboflavin, nicotinic acid, pantothenic acid, p-aminobenzoic acid,

folic acid, lipoic acid, B12, etc.

c Others Purine bases, pyrimidine bases, choline, inositol, peptides, etc.

a Where growth proceeds in the absence of growth factors, the compounds are presumably synthesized by the organism.

Source: Alexander, M Introduction to Soil Microbiology 2nd ed John Wiley & Sons, New York 1977 With permission.

carbon dioxide Thus, efficiency of conversion of substrate (petroleum) carbon to cellular material isless than 100% The optimal C:N and C:P ratios are expected to be wider than the theoretical values.

It has been suggested that a C:N ratio of <25 leads to mineralization (excessive N present) and aC:N ratio of >38 leads to depletion of mineralized N (Routson and Wildung, 1970) The latter conditionwould limit biodegradation due to nitrogen starvation

The ratio depends upon the rate and extent of degradation of the chemicals involved and may varyaccording to the particular contaminants present Biodegradation of complex oily sludges in soil occursmost rapidly when nitrogen is added to reduce the C:N ratio to 9:1 (Brown, Donnelly, and Deuel, 1983b).That of petrochemical sludge is most rapid when nitrogen, phosphorus, and potassium are added at arate of 124:1, C:NPK The optimal ratios may be different for different soils (Zhou and Crawford, 1995).These authors found the optimal C:N ratio for a clay/loam soil to be around 50:1 Dibble and Bartha(1979) reported a C:N ratio of 60:1 and a C:P ratio of 800:1 On the other hand, excessive nitrogen(e.g., C:N = 1.8:1) can impair biodegradation, possibly due to ammonia toxicity, although a greateramount of fertilizer has not been shown to inhibit biodegradation (Zhou and Crawford, 1995).During experiments on landfarming waste oil, it was determined that carbon-to-nitrogen and carbon-to-phosphorus ratios of 60:1 and 800:1, respectively, were optimal under the conditions used (Dibbleand Bartha, 1979a) Addition of yeast extract or domestic sewage did not prove beneficial Urea form-aldehyde was found to be the most satisfactory nitrogen source tested, since it effectively stimulatedbiodegradation and did not leach nitrogen, which could contaminate the groundwater (Dibble and Bartha,1979b) A problem with this technology is that runoff water from the site could contain high amounts

of oil and fertilizer (Kincannon, 1972)

Under most growth conditions, about half of the carbon available from growth hydrocarbons tually becomes cellular biomass (Texas Research Institute, Inc., 1982) This consists primarily of proteins,nucleic acids, amino acids, purines, pyrimidines, lipids, and polysaccharides Huesemann and Moore(1993) observed an 11-fold increase in polar nonhydrocarbon compounds after 22 weeks of biodegra-dation of crude oil–contaminated soil The initial amount of 430 mg/kg rose to 4725 mg/kg The increase

even-in polar compounds was probably due to biomass (cell mass) formation, citeven-ing, as above, constituentssuch as intracellular proteins, lipids, carbohydrates, and material found in cell walls and membranes

Growth of Rhodococcus and Pseudomonas strains on naphthalene, fluorene, phenanthrene, anthracene,

fluoranthene, and pyrene yielded high mineralization (56 to 77% of the carbon) and good production ofbiomass (16 to 35% of carbon) and limited, but significant, accumulation of metabolites (5% to 23% ofcarbon) (Bouchez, Blanchet, and Vandecasteele, 1996) The higher PAHs produced higher mineralizationand lower biomass

The nitrogen requirement value (NRV) is the amount of nitrogen required by microorganisms todecompose/degrade a particular organic chemical waste (Parr, Sikora, and Burge, 1983) It dependsmainly upon two factors: the chemical composition of the waste and the rate of decomposition Thisvalue is also affected by the other soil factors

After contact with an oily waste, microbial activity initially decreases (Hornick, Fisher, and Paolini,1983) This may be due to the same initial decrease in mineral nitrogen resulting from nitrogen immo-bilization by hydrocarbon-metabolizing microbes using up all the available nitrogen In time, the micro-organisms will adapt to the high C:N ratio and increase the total microbial population (Overcash andPal, 1979a)

Addition of sodium nitrate enhanced the oxidation of hydrocarbons and the ultimate decay of theresulting organic carbon compounds to inorganic carbon compounds (Dietz, 1980) Addition of potassiumorthophosphates, KH2PO4 and K2HPO4, had no effect on biodegradation in this application The phos-phate precipitated in a very early stage due to the presence of calcium In another case, mineralization

of phenanthrene was found to be enhanced by addition of phosphate but not potassium, while it wasreduced by addition of nitrate (Manilal and Alexander, 1991)

Calcium promotes flocculation, the clumping of tiny soil particulates, and may prevent thoroughincorporation of phosphate into the soil (Brady, 1974) U.S EPA (1985a) also reported that addition ofphosphates can result in the precipitation of calcium and iron phosphates At low concentrations incalcareous soils, phosphorus is adsorbed onto calcium carbonate; at high concentrations, calcium phos-phate minerals are formed, and the phosphate is not available to the microorganisms (Mattingly, 1975).Phosphate can be added to sandy (quartz) soils, but not to calcareous soils (Aggarwal, Means, andHinchee, 1991)

If calcium is present at 200 mg/L, it is likely that calcium supplementation is unnecessary (U.S EPA,1985a) Calcium deficiencies usually occur only in acid soils and can be corrected by liming (JRBAssociates, Inc., 1984) If the soil is deficient in magnesium, the use of dolomitic lime is advised It isdesirable to have a high level of exchangeable bases (calcium, magnesium, sodium, and potassium) onthe surface exchange sites of the soil for good microbial activity and for preventing excessively acidconditions Sulfur levels in soils are usually sufficient; however, sulfur is also a constituent of mostinorganic fertilizers.

The common form of phosphorus in soil is H2PO4–1in basic soil solutions (Mattingly, 1975) phorus concentrations in the soil solution are usually low, ranging from 0.1 to 1 ppm, since this element

Phos-is mostly associated with the solid phase in soils (Hornick, 1983) In acid soils, phosphorus reacts withiron and aluminum hydroxides to produce adsorbed forms of phosphorus that are in equilibrium withthe soil solution or are precipitated and, thus, occluded by the minerals (Dietz, 1980) Robertson and

Alexander (1992) report that the extent of growth of a Corynebacterium sp was reduced with 2 or

10 mM phosphate in media containing high iron concentrations

Phosphorus concentration affects the pH at which mineralization can occur (Robertson and Alexander,

1992) A Pseudomonas sp mineralized phenol rapidly in medium with 0.2 mM phosphate at pH 5.2,

but had little or no effect at pH 8.0 However, mineralization proved to be greater at pH 8.0 than at pH5.2, when the culture contained 10 mM phosphate

Microorganisms may be limited by phosphorus but not nitrogen (Thorn and Ventullo, 1986) Neithernitrogen nor phosphorus enrichment alone stimulated the biodegradation of phenol in topsoil (Atlas andBartha, 1973d) However, in two different types of subsurface soils, addition of these nutrients signifi-cantly stimulated mineralization Phosphorus enrichment had the greatest effects, and the effects ofsimultaneous nitrogen and phosphorus amendments were similar to those observed with phosphorusalone Phosphorus limitation may be widespread in subsurface soils If the input of phosphorus into thesubsurface is disproportionate to that of organic compounds, phosphorus limitation could greatly reducethe ability of microbes in deeper soil to degrade pollutants as they migrate downward, assuming oxygen

or another electron acceptor is not limiting

Huesemann and Moore (1993) determined that nitrogen and phosphorus fertilizers were importantfor stimulating biodegradation of petroleum hydrocarbon–contaminated soils, while addition of bacteriafrom activated sludge solids alone was not sufficient Degradation by addition of cow manure alone wasequivalent to that of the fertilizer over the span of a year; however, the biodegradation rate was optimalwith a mixture of the fertilizer, an activated sludge inoculum, and cow manure

Ammonium phosphate generally provides the nitrogen and phosphorus required for maximum growth

of hydrocarbon oxidizers (Rosenberg, Legmann, Kushmaro, Taube, Adler, and Ron, 1992) A mixture

of other salts, such as ammonium sulfate, ammonium nitrate, ammonium chloride, sodium phosphate,potassium phosphate, and calcium phosphate, can also be used However, these are all highly watersoluble and can become too dilute in the environment to maintain their effectiveness Oleophilic nitrogenand phosphorus compounds with low C:N and C:P ratios can overcome this problem

Fixed nitrogen can initially be a limiting nutrient, but nitrogen limitation can sometimes be overcome

by nitrogen fixation (Toccalino, Johnson, and Boone, 1993) This was observed in butane-amended soilbut not in propane-amended soil

The main danger at hazardous waste sites may be in overloading the soil with elements that mayhave been present in the waste (or already in the soil, e.g., phosphate plugging), causing toxicity andleaching problems (JRB Associates, Inc., 1984) Nutrient formulations often contain excessive ortho-phosphate to decrease the rate of peroxide decomposition (Aggarwal, Means, and Hinchee, 1991) Themaximum orthophosphate concentration that may provide microbial nutrients and yet avoid significantprecipitation in most geochemical environments is about 10 mg/L (Miller and Hinchee, 1990) In order

to achieve this concentration at a greater distance, however, higher concentrations must be introduced

at the injection point, where excessive precipitation may result Thus, the plugging problem may beunavoidable with orthophosphate-based nutrient formulations An available soil phosphate content ofabout 20 mg/L may be needed for microbial growth (Aggarwal, Means, Hinchee, Headington, Gavaskar,Scowden, Arthur, Evers, and Bigelow, 1990)

Polyphosphates (e.g., pyrophosphate, tripolyphosphate, and trimetaphosphate) can serve as an native source of phosphate to avoid plugging problems (Aggarwal, Means, and Hinchee, 1991) Restore

alter-375 incorporates sodium tripolyphosphate, but in combination with orthophosphates Polyphosphate

hydrolysis is influenced by pH, ionic composition of the solution, microbial activity, concentration ofthe enzyme polyphosphatase, and chain length (Gilliam and Sample, 1968; Blanchar and Hossner, 1969;Blanchar and Riego, 1976; Busman and Tabatabai, 1985; Dick and Tabatabai, 1986; Hons, Stewart, and

Hossner, 1986) Polyphosphates have a half-life of 1 to 10 days and can be used as in situ, slow-release

sources of orthophosphate Another polyphosphate that has been successfully applied at a fueloil–contaminated site is sodium hexametaphosphate (Steiof and Dott, 1995) Poly- and metaphosphatesboth have complexing qualities for cations such as Ca2+ and Mg2+ and thus do not precipitate easily withthese cations They interact with soil, but their sorption is not strong, so they can be transported overlonger distances, with metaphosphate being better for this purpose

Influent substrate-loading rates directly affect the injection pressure and, thus, near-well biofouling(Jennings, Petersen, Skeen, Peyton, Hooker, Johnstone, and Yonge, 1995) Substrate-loading rates alsodetermine biomass concentration in effluents Continuous nutrient addition to the soil causes biomass

to concentrate near the nutrient injection point

Both total biomass production and growth efficiency are inversely correlated with EPS production,which can lead to subsurface plugging, under aerobic or anaerobic conditions (Hanneman, Johnstone,Yonge, Petersen, Peyton, and Skeen, 1995) The oxidation state of the carbon source can be used toestimate the potential for EPS production Substrates more reduced than glucose yield more energy thatcan be used in synthesis of polysaccharide; substrates more oxidized than glucose allow energy to beused for cell synthesis rather than for producing large quantities of EPS The type of cometabolite (carbonsource) used to stimulate biomass production under denitrifying conditions will influence the amount

of EPS production, which could lead to subsurface plugging

Temperature also plays a role when nutrients are added (Walworth and Reynolds, 1995) At 10°C,bioremediation rates are not affected by addition of phosphorus or nitrogen; however, at 20°C, biore-mediation is increased by addition of phosphorus but not nitrogen

Key trace elements are essential to the stimulation of bacterial growth (Kincannon, 1972) Thesemicronutrients are required in such small doses that most are already abundant in the soil They includesulfur, sodium, calcium, magnesium, and iron Copper, zinc, and lead are normally considered to exhibitharmful effects on biological growth Addition of yeast cells can serve as a nutrient source (Lehtomakeiand Niemela, 1975) Organic and inorganic nutrients in natural waters affect the rate of mineralization

of organic compounds in trace concentrations (Kaufman and Doyle, 1978) Inorganic nutrients, arginine,

or yeast extract often enhance, but glucose reduces, the rate of mineralization

The concentration of nutrients and organics should be kept as uniform as possible to protect againstshock loading (U.S EPA, 1985a) Nitrogen must be applied with caution to avoid excessive application(Saxena and Bartha, 1983) Nitrate or other forms of nitrogen oxidized to nitrate in the soil may beleached to the groundwater (nitrate is itself a pollutant limited to 10 mg/L in drinking water) (U.S EPA,1985a) Some nitrogen fertilizers may also tend to lower the soil pH, necessitating a liming program tomaintain the optimal pH for biological activity Since the pH generally decreases with growth with theuse of ammonium salts of strong acids, urea can serve as a nitrogen source (Rosenberg, Legmann,Kushmaro, Taube, Adler, and Ron, 1992) Low concentrations of readily metabolized organic compounds(peptone, calcium lactate, yeast extract, nicotinamide, riboflavin, pyridoxine, thiamine, ascorbic acid)often promote the growth of the oxidizer, but high concentrations will retard the degradation of thehydrocarbons (ZoBell, 1946; Morozov and Nikolayov, 1978) The quantity of organic material to addmust be determined in treatability studies (Sims and Bass, 1984) Nutrient formulations should be devisedwith the help of experienced geochemists to minimize problems with precipitation and dispersion ofclays (U.S EPA, 1985a) Special soil preconditioners and nutrient formulations to reduce these problemsand maximize nutrient mobility and solubility are being investigated

Population turnover allows for the recycling of nutrients (Dibble and Bartha, 1979a) However, it isexpected that fertilizer in the optimal ratios will have to be reapplied, as necessary The best fertilizersfor soil application are in a form of readily usable nitrogen and phosphorus and also in a slow-releaseform to provide a continuous supply of nutrients, which is beneficial in terms of fertilizer savings andminimized leaching from the oil–soil interface (Atlas, 1977) A liquid fertilizer containing: 3340 lbammonium sulfate, 920 lb disodium phosphate, and 740 lb monosodium phosphate was injected intowells at a contaminated site in Marcus Hook, PA (Raymond, Jamison, and Hudson, 1976) Addition ofnutrients in this form accelerated the removal of contaminating gasoline

An additive has been developed to promote biodegradation of materials such as hydrocarbons andoil spills (Basseres, Eyraud, and Ladousse, 1994) The additive consists of a mixture of at least oneassimilable nitrogen source, composed of at least one unsubstituted or substituted amino acid, and atleast one phosphorus source, e.g., phosphate rocks, with the ratio of nitrogen to phosphorus rangingfrom 2 to 100 Meat or fish meal is acylated with lauryl acid chloride to render the additive oleophilicand mixed with an amino acid, e.g., lysine, methionine, cystine, threonine, tryptophan, hydroxylysine,

or hydroxyproline The composition is an effective nutrient and has been found to allow biodegradation

of oil spills on ocean water

Kopp-Holtwiesche, Weiss, and Boehme (1993) have patented an improved nutrient mixture forbioremediation of polluted soils and waters The nutrient mixture to enhance biodegradation of pollutants,especially hydrocarbons, contains phosphoric acid ester emulsifiers as a phosphorus source and one ormore water-soluble or dispersible nitrogen sources It may also contain biodegradable surfactants, withthe exception of glycerin esters Nonionic surfactants are preferred Suitable phosphoric acid estersinclude glycerophospholipids (Lipotin), other phospholipids, alkyl phosphates, and/or alkyl-ether phos-phates This composition is useful for biodegradation of oil spills on soils

There are many substances that would be suitable as fertilizers, and their compositions and originsdiffer considerably (Sims and Bass, 1984) The choice of an appropriate fertilizer can be complicated,and an agronomist should be consulted to develop a fertilization plan at a hazardous waste site A planmay include types and amounts of nutrients, timing and frequency of application, and method ofapplication The nutrient status of the soil and the nutrient content of the wastes must be determined toformulate an appropriate fertilization plan

An optimum fertilization program has been proposed (Kincannon, 1972) Chemicals are added so as

to attain a slight excess of nitrogen, phosphorus, and potassium in the contaminated area After that, soiltesting for ammonia and nitrates is conducted at regular monthly intervals Small doses of ammoniumnitrate are added, as needed, to maintain the ammonium or nitrate surplus Urea is used as a nitrogensource to avoid the initial increase in soil salts, which may result from additions of other fertilizer stocks.Ammonium nitrate is subsequently applied, once urea is deemed no longer necessary Potash is added

Vented percolation in situ can be used to stimulate naturally occurring organisms to degrade

hydro-carbons by supplying nutrients and oxygen to the subsurface (Ram, Bass, Falotico, and Leahy, 1993).After oxygen is introduced by a vacuum-inducing airflow through the soil, nutrients can be percolatedthrough the soil with the vent-system piping Volatile hydrocarbons are removed by the venting, leavingadsorbed, heavier hydrocarbons to be biodegraded Water (recovered, treated groundwater or freshwater)

is amended with nutrients and injected under supplied or gravity pressure to the vent system with theblower off The nutrients could otherwise be dispersed through horizontal slotted piping laid at intervals

on the surface or in ditches just above the depth of contamination

The BioPurgeSM technology is based on the low-volume airflow of bioventing, but with a closed-loopsystem (Burke and Rhodes, 1995) Soil vapor is extracted from wells in the contaminated soil and treated

in an ex situ unit, where the volatiles are absorbed and biodegraded Oxygen, nutrients, heat, and moisture

are added to the vapor stream, which is reinjected into the contaminated soil above the water table Theadditives can be monitored, and there are no off-gases to treat This approach has limited success in siltsand clays See Section 2.2.2.4 for a full description of the process

Deep Soil Fracture Bioinjection™ employs pressurized subsurface injection to introduce release nutrients, oxygen, and microbes, which permeate all types of soil to ensure excellent transmittance

controlled-(Burke and Rhodes, 1995) This is an effective and economical technique, which is described in

contam-Widrig and Manning (1995) determined that continuous saturation with nitrogen and phosphorusamendments was not as effective as periodic operation, consisting of flooding with nutrients, followed

by draining and forced aeration By monitoring CO2 and O2 levels in situ, it may be possible to optimize

the timing of flooding and aeration events to maximize degradation rates

Gaseous ammonia can be introduced with bioventing techniques to the subsurface at sites wheresurface application of liquid nutrients is not possible (Marshall, 1995) The ammonium ion is the preferrednutrient form of many microorganisms Ammonia supplied to the subsurface dissolves readily in soilmoisture and sorbs strongly to soil particles Such ammonia applications should be conservative toenhance biodegradation Ammonia-oxidizing organisms will convert some of the ammonia to nitrate,and excessive ammonia can promote formation of methane from anaerobic hydrocarbon degradationwith nitrate as the electron acceptor

Dineen, Slater, Hicks, Holland, and Clendening (1993) add anhydrous ammonia as a source of reducednitrogen to an airstream through the unsaturated soil This increases the soil oxygen and nitrogen levels,resulting in a 100-fold increase in microbial count Maintaining viable cell counts at the level of 106 to

107 should result in a decrease of petroleum hydrocarbons in situ to a cleanup level of 100 ppm.

A microcapsule technique has been patented for degrading hydrocarbons (Schlaemus, Marshall,MacNaughton, Alexander, and Scott, 1994) It consists of a core material, a coating material, and atleast one microorganism capable of degrading the hydrocarbon The core is lipophilic material containingnutrients for the microorganisms The coating material is water soluble The capsule is such that theorganisms are kept in close proximity to the hydrocarbon to be degraded

Another encapsulation approach employs liposomes (Gatt, Bercovier, and Barenholz, 1991) Thephospholipids in liposomes are naturally occurring membrane lipids (e.g., plant phospholipids), whichcan serve as a source of carbon, hydrogen, phosphorus, and nitrogen for microorganisms in the subsur-face The liposomes are in the shape of sealed microsacs containing water, which can provide a reservoirfor nutrients, minerals, sugars, amino acids, vitamins, hormones, drugs, and growth factors The mem-brane of the vesicle is hydrophobic on the inside and hydrophilic on the outside, which makes itcompatible with a variety of neutral or charged, lipophilic or amphiphilic compounds Continuous release

of the nutrients could be controlled by the lipid composition Liposomes rapidly induced a considerableenhancement (up to 7 logs) of growth of bacteria in soil contaminated with petroleum Counts of over

1011 cells/mL were obtained The liposomes could be used to clean up contaminated soils both byenhancing microbial growth and by modifying the physical properties of the contaminant (reducing theinterfacial tension 10,000- to 50,000-fold), making it more biodegradable (See Section 5.3.1.2) Thereare numerous ways they could be dispersed, and they should have no toxic effects on the environment.Liposomes can be prepared on a small or large scale and be tailor-made to suit the requirements of thespecific microorganisms and chemicals involved

A similar patent involves a carrier for supporting microorganisms for soil remediation (Kozaki, Kato,Tanaka, Yano, Sakuranaga, and Imamura, 1994) This carrier contains pores, which hold a nutrient orare a nutrient for the organisms A method for remediating the soil by administering the carrier has beendeveloped

In most cases, site geology will determine the method of fertilization to be used in a given situation(Raymond, Jamison, and Hudson, 1976) For example, in a fractured dolomite and clay formation, lack

of homogeneity makes well injection and distribution of nutrients difficult Use of diammonium phate could result in excessive precipitation, and nutrient solution containing sodium could causedispersion of the clays, thereby reducing permeability (U.S EPA, 1985a) High calcium could causeprecipitation of added phosphate, rendering it unavailable to microbial metabolism If a site is likely toencounter problems with precipitation, iron and manganese addition may not be desirable If the totaldissolved solids content in the water is extremely high, it may be desirable to add as little extra salts aspossible

phos-Results of oil biodegradation in Marcus Hook, PA, and Corpus Cristi, TX, indicated that fertilizerwas not a factor in biodegradation until approximately 50% of the oil had been degraded (Raymond,Hudson, and Jamison, 1976) This cannot be regarded as conclusive, since other environmental factorsmay have affected these studies.

A large kerosene spill (1.9 million L) in New Jersey was cleaned up by a combination of techniques(Dibble and Bartha, 1979c) Much of the kerosene was recovered by physical means and by removing

200 m3 of contaminated soil Following stimulation of microbial degradation by liming, fertilization,and tillage, phytotoxicity was reduced

Addition of nitrogen and phosphorus fertilizer at another site resulted in a doubling of the oilbiodegradation rate of 70 bbl/acre/month to 1.0 lb/ft3/month (Kincannon, 1982) It is recommended thatmonthly determinations of nitrogen and phosphorus levels in the soil and periodic fertilizer application,when necessary, will optimize the fertilization process The cost of soil disposal of oily wastes wasestimated at $3.00/bbl Degradation rates of up to 100 bbl/acre/month were reported, when the oil wasapplied to fertilized soils (Francke and Clark, 1974)

In the 1989 Exxon Valdez oil spill at Prince William Sound, AK, the rate of natural degradation waslow and limited by environmental factors (Pritchard, Mueller, Rogers, Kremer, and Glaser, 1992; Atlas,1995) This oil spill provided an opportunity for a major study of the effect of fertilizers on bioremedi-ation The efficiency of a fertilizer depends greatly on the environment and design of the treatmentprotocol Hydrocarbon degraders are normally less than 1% of the total microbial community Thisincreases to about 10%, when oil pollutants are present These are the organisms that need to be stimulated

by providing the appropriate nutrients

Fertilizers were able to enhance biodegradation of the indigenous hydrocarbon degraders in thiscleanup effort (Pritchard, Mueller, Rogers, Kremer, and Glaser, 1992; Atlas, 1995) Three types ofnutrient supplementation were used One was a water-soluble (23:2 N:P) garden fertilizer formulation.Another was Customblen, a slow-release calcium phosphate, ammonium phosphate, and ammoniumnitrate within a polymerized vegetable oil coating The third was the oleophilic fertilizer Inipol EAP-22(developed by Elf Aquitaine in France), an oil-in-water microemulsion with urea as a nitrogen source,tri(laureth-4)phosphate as a surfactant and a phosphorus source (an N:P ratio of 7.3:2.8; Glaser, 1991),2-butoxyl-1-ethanol, and oleic acid as a carbon source Multiple regression models showed nitrogenapplications were effective in stimulating biodegradation rates The failures and successes were discussed

by these authors, as well as the necessary prerequisites that must be met for fertilizers to work

A problem with Inipol EAP 22 is that it contains a large amount of oleic acid, which increases theC:N ratio and can serve as an alternative carbon source for the organisms (Rosenberg, Legmann,Kushmaro, Taube, Adler, and Ron, 1992) It also contains an emulsifier, and contact with water releasesthe urea to the water phase, where it is not available for the microorganisms

A new, controlled-release, hydrophobic fertilizer, F-1, has been developed to overcome many of theproblems associated with other sources of nitrogen and phosphorus (Rosenberg, Legmann, Kushmaro,Taube, Adler, and Ron, 1992) F-1 is a modified urea-formaldehyde polymer containing 18% N and10% P as P2O5 It is insoluble and attaches to the oil/water interface with the microorganisms Strains

with a cell-bound, inducible enzyme for F-1 (e.g., Pseudomonas sp., Gluconbacter strain RT, nas strain RL4, and P alcaligenes strain RL3) can depolymerize the fertilizer to obtain nutrients at the

Pseudomo-site where they are needed These microbes with high cell–surface hydrophobicity then desorb from thehydrocarbon when it is depleted, by releasing a hydrophilic capsular material, which repels them fromthe spent droplet and allows them to reattach to a fresh drop (Rosenberg, 1986; Rosenberg, Rosenberg,Shohan, Kaplan, and Sar, 1989) The fertilizer can continue to support growth and biodegradation, evenafter the aqueous phase is removed (Rosenberg, Legmann, Kushmaro, Taube, Adler, and Ron, 1992) Amixed culture of bacteria containing F-1-ase and growing on crude oil reached 1 × 108 cells/mL F-1-ase activity is associated with the cells rather than the extracellular fluid, and it is influenced by thesource of nitrogen and phosphorus

Degradation of Nigerian light crude oil by Bacillus strains 28A and 61B was enhanced by addition

of organic nitrogen sources (0.7% peptone, 0.14% urea, and 0.7% yeast extract), while inorganic nitrogensources (0.46% KNO3 and 0.3% (NH4)2SO4) had a depressing effect on the degradation

In the coral-derived sands of Kwajalein Island, in the Republic of the Marshall Islands, bioremediation

of diesel fuel–contaminated soil by indigenous organisms was found to be feasible, but the degradationrates were very low (Siegrist, Phelps, Korte, and Pickering, 1994) The sand was alkaline (pH > 8) and

deficient in nutrients (low nitrogen and phosphorus) Addition of nutrients enhanced the degradationsomewhat.

Biodegradation is not always stimulated by addition of inorganic nutrients, because other factors maysuppress microbial activity or interact with nutrient limitation to slow degradation (Steffensen andAlexander, 1995) The presence of other bacteria or other substrates may reduce the degradation ofcertain compounds, possibly as a result of competition for the nutrients

Organic material is very important in the soil matrix (Hornick, 1983) The presence of organic materialsmay have many effects on soil properties, including soil structure, water-holding capacity, bulk density,mobilization of nutrients (hindering degradation of organic wastes), reduction in soil erosion, and soiltemperature (Atlas, 1978a)

Naturally occurring organic material can influence the ability of microorganisms to degrade pollutants(Shimp and Pfaender, 1984) Its role in metal reactions or sorption processes that occur in the soildetermines the availability of metals and essential nutrients for plants and microorganisms (Hornick,1983) Sorption of contaminants on soil particles can alter the molecular character and enzymatic attack

of a given compound

Soil generally contains 5 to 12% organic matter (Overcash and Pal, 1979a), and, of that, some 1 to4% is mineralized annually (McGill, 1980) Assuming that 5% of the organic matter is nitrogen and 0.5%

is phosphate, some 50 to 480 lb/acre nitrogen (60 to 550 kg/ha) and 5 to 48 lb/acre phosphorus (6 to

55 kg/ha) are available annually from soil for use by soil microflora to grow on waste added to the soil.Soil contains organic material in varying stages of decomposition (JRB Associates, Inc., 1984).Organic matter is generally an amorphous organic residual in soils, which, when present in sufficientamounts, has a beneficial effect on the physical and chemical properties of the soil (Hornick, 1983).Around 65 to 75% of the organic material in soil (60 to 80% in most groundwaters and sediments; Khan,1980) usually consists of humic substances, i.e., humic acid, fulvic acid, and humin (Schnitzer, 1978;Hornick, 1983) These humic substances have very large surface areas, large amounts of exchangeablebases, and high cation exchange capacities (the total amount of cations held exchangeably by a unitmass or weight of a soil) The crude humin consists of humic acid and hymatomelanic acids containingfunctional carboxyl and phenolic hydroxyl groups responsible for exchange and adsorption reactions.Both humates and fulvates show a high degree of reactivity due to their acidic functional groups Thereaction of these materials with cations in the soil solution is strongly pH dependent These organicstend to be recalcitrant to degradation (Khan, 1980) The remainder of the organic material consists ofpolysaccharides and proteins, such as carbohydrates, proteins, peptides, amino acids, fats, waxes, alkanes,and low-molecular-weight organic acids, which are rapidly decomposed by the soil microorganisms(Schnitzer, 1982) This organic matter can also contribute nitrogen, phosphorus, sulfur, zinc, and boron,all of which add to the nutrient status of the soil (JRB Associates, Inc., 1984) Easily decomposed organiccontaminants can become part of an important soil process and result in a substantial increase in beneficialorganic materials (Hornick, 1983) It is likely that maintaining a supply of biodegradable organic matter

in site soils would allow a higher population of diverse microbes capable of degrading many kinds oftoxic organic compounds

Organic matter is very important to the microbial ecology and activity of the soil (Sims and Bass,1984; JRB Associates, Inc., 1984) Its high cation exchange capacity and high density of reactivefunctional groups help to bind both organic and inorganic compounds that may be added to the soil.These properties also help to retain the soil bacteria which can then attack the bound compounds Thus,the sorbents may immobilize the organic constituents, as well as allow more time for biodegradation.Bacteria seem to be able to survive better in the presence of organic matter, especially in drier soils(Godbout, Comeau, and Greer, 1995) The presence of the solid organic matrix appears to be essentialfor enhanced degradation (Kaestner and Mahro, 1996) Humus increases the water-holding capacity ofsoil by swelling when wet to absorb two to three times its weight in water (Hornick, 1983) Because ofits surface area, surface properties, and functional groups, humified soil can serve as a buffer, an ionexchanger, a surfactant, a chelating agent, and a general sorbent to help in the attenuation of hazardouscompounds in soils (Ahlrichs, 1972)

On the one hand, humic polymers act as stabilizing agents, making contaminants less resistant tobiodegradation (Verma, Martin, and Haider, 1975), while, on the other hand, compounds bound to humic

material can become unavailable for biodegradation Manilal and Alexander (1991) suggest that radation of PAHs might be slowed by their sorption to soil organic matter After adaptation of a microbialcommunity to four types of compounds, it was found that amino acids, fatty acids, and carbohydratesstimulated biodegradation of monosubstituted phenols, while humics decreased biodegradation rates(Shimp and Pfaender, 1984) It appears that fulvic acids may be toxic to microbes and cause sorption

biodeg-of a contaminant, making it less bioavailable (Grosser, Warshawsky, and Kinkle, 1994)

Enzyme activities of soil organisms can be responsible for catalyzing the binding of xenobioticcompounds and their breakdown products to soil humic materials (de Klerk and van der Linden, 1974;Bollag, 1983) Bound hazardous organic compounds, including toxic metabolites, should be monitored.Humus-bound xenobiotic compounds may be slow to mineralize or be transformed to innocuous forms(Khan, 1982) In these cases, the humic content of the soil should probably not be increased Hazardousconstituents may be initially bound to organic materials, but later released as organic materials decompose(Sims and Bass, 1984) The released materials may be subject to leaching, volatilization, or reattachment

to soil organic matter This suggests that treatment is not complete until it can be demonstrated thatthese compounds are absent or at a safe level in the soil (Bollag, 1983)

In waste-amended soils, the addition of high amounts of organic matter ensures a predominance oforganic matter reactions (Schnitzer and Khan, 1978) The mobility of heavy metals added by wastes isrelated to the organic matter content of soils, pH, hydrous oxide reactions, and the oxidation-reduction

or redox potential of a soil If the soil contains cracks and fractures that may increase the potential formobilization and groundwater contamination, addition of an adsorbent can be useful (Sims and Bass,1984; JRB Associates, Inc., 1984) It is especially important and effective in soils with low organicmatter content, such as sandy and strip-mined soils These sorbants include agricultural products andby-products, sewage sludges, other organic matter, and activated carbon

Organic matter has a beneficial effect when bacteria are added immediately after soil contamination(Godbout, Comeau, and Greer, 1995) However, if the bacteria are added after 38 days pre-exposure tothe chemicals, this beneficial effect is observed only in sand The negative effect of soil texture oncontaminant mineralization is more significant with time, possibly due to the formation of clay–humicacid complexes that increase the adsorption of substrate and nutrients on soil particles making them lessbioavailable (Godbout, Comeau, and Greer, 1995)

Supplemental carbon and energy sources can be used to stimulate the metabolism of even recalcitrantxenobiotics, either through cometabolism (Alexander, 1981) or simply because of the presence of additionalcarbon and energy (Yagi and Sudo, 1980) However, if biodegradable organic materials are added to thesoil in order to raise the C:N ratio higher than about 20:1, mineral nitrogen in the soil will be immobilizedinto microbial biomass, and the decomposition process will be slowed considerably (JRB Associates, Inc.,1984) Phosphorus is similarly immobilized when carbon is in excess (Alexander, 1977) If the soil must

be managed to decompose organic matter during the treatment of hazardous waste–contaminated soils,nitrogen and phosphorus may be required to bring the C:N:P ratio close to that of the bacterial biomass.Terrestrial oil spillages will probably result in the death of plants, releasing large amounts of nonhydrocarbonorganic matter into soil, which can serve as an alternative source of carbon for heterotrophic microorgan-isms, thereby interfering with the degradation of the contaminants (Atlas, 1977)

Natural organic matter can be added to the soil, such as in the form of synthetic commercial organics,cattle manure, sewage sludge, or crop residues (Arthur D Little, Inc., 1976) Commercial syntheticorganics are expensive and their suitability for microbial growth is uncertain

Mixed results have been obtained by different researchers with using manure amendments to increasethe rate of degradation of organic chemicals While some workers reported that manure amendmentsincreased the rate of degradation of ten organic chemicals tested, Doyle (1979) found that manure didnot significantly reduce the degradation of any chemical examined The breakdown of several compoundswas positively correlated with the increased total microbial activity of manure-amended soil Sewagesludge, however, enhanced the breakdown of only two compounds, while decreasing the rate of degra-dation of nine others Some advantages of using municipal sludges in organic waste treatment are thatthey contain active indigenous populations of microorganisms with degradative potential, and theyprovide necessary nutrients for biodegradation (Sims and Bass, 1984)

Sewage sludge and cattle manure are the least-expensive supplements; however, their use is limitedsince they contain variable quantities of trace elements or heavy metals that may disturb the expected