• Chromium;
  • Lead;
  • Speciation;
  • Bioavailability;
  • Ecological risk assessment


  1. Top of page
  2. Abstract
  7. Acknowledgements

When evaluating the risk chemicals may pose to mammals and birds in ecological risk assessments (ERAs), it is common practice to conservatively assume that all (100%) of a chemical in an environmental medium is bioavailable to receptors. This assumption often leads to overestimating ecological risk and may ultimately result in costly and unnecessary risk management actions. While effects of bioavailability and speciation of metals such as arsenic (As) and lead (Pb) have been considered in human health risk assessment, these effects are rarely taken into consideration when assessing risks to mammals and birds. An ERA was conducted at the former Col-Tex refinery site in Colorado City, Texas, USA, to characterize risks to select wildlife species from exposure to chromium (Cr) and Pb found in soils. The focus on these metals was based on results of a screening-level ERA that found that Cr and Pb were posing ecological risks at the site. Soils were analyzed for total Cr and Pb, trivalent Cr (CrIII), hexavalent Cr (CrVI), organic Pb, and the bioavailability and speciation of Pb. Results for Pb and Cr indicated that >94% of the Cr was present as the less toxic and immobile Cr(lll) and that >99% of the Pb in soils was present as inorganic Pb. Lead bioaccessibility measured by in vitro testing ranged from 8% to 77.8%, depending on location of individual soil samples. Results demonstrated that Pb and Cr bioavailability and speciation information can raise soil cleanup concentrations while being protective of ecological receptors. The costs of performing the ERA were de minimus compared to the reduction in remediation costs at the site. The refined hazard estimates allowed informed decision making in the management and segregation of soils, allowing for effective risk management at the site.


  1. Top of page
  2. Abstract
  7. Acknowledgements

When evaluating the risk of chemicals in screening-level ecological risk assessments (ERAs), it is common practice to use conservative assumptions regarding bioavailability and toxicity. For example, it is often assumed that a contaminant in an environmental medium such as soil is 100% bioavailable to a receptor. Similarly, the toxicity of a contaminant is often derived from laboratory studies that use soluble forms of the contaminant that may not be present in the medium under consideration. In the case of lead (Pb), toxicity values for mammals and birds are often derived from oral exposures to highly soluble Pb acetate (USEPA 1997a; NRC 2003; Schoof 2003a; USEPA 2005). Lead acetate is more readily absorbed from the gastrointestinal (GI) tract than other lead compounds, thus maximizing bioavailability (Chaney et al. 1989; Freeman et al. 1992; Ruby et al. 1992; Davis et al. 1994), and 100% bioavailability is assumed. But for many contaminants assessed in this manner, there are substantial differences in the amounts that are bioavailable from different media (Chaney et al. 1989; Dieter et al. 1993; Schoof et al. 1995: Casteel et al. 1997). Ignoring these differences in bioavailability can lead to an overestimation of risk to ecological receptors by assuming that exposure to the chemical form in ambient media does not differ from contaminants administered in laboratory media. The oral toxicity threshold values for Pb and chromium (Cr) used in screening-level ERAs generally are derived from literature of laboratory studies of soluble salts of these metals in diet and drinking water (Schroeder et al. 1963; Grant et al. 1980; Kimmel et al. 1980; Edens and Garlich 1983). However, in contrast to the highly soluble Pb in drinking water or diet used in laboratory studies, Pb and Cr in soils generally exist in mineral forms that would not be completely solubilized in the GI tract (Witmer et al. 1991; Freeman et al. 1992, 1994, 1996; Ruby et al. 1992). For metals such as Pb and Cr to be absorbed, they must 1st be dissolved (Freeman et al. 1992; Hamel et al. 1998). Thus, Pb or Cr in soil will be less absorbed than metals in drinking water or diet without soil (Witmer et al. 1991; Freeman et al. 1992, 1996; Ruby et al. 1996). For example, the addition of 5% uncontaminated soil to the diet of rats reduced the bioavailability of Pb acetate by 47% as measured by Pb concentrations in tibia (Chaney et al. 1989). Unless this reduction in relative bioavailability of the metal is taken into consideration, risks to terrestrial receptors are likely to be overestimated in further refinements of the ERA. Accepting an uncertain and overstated risk estimate may result in costly and unnecessary risk management actions. Site-specific measurements of metals' relative bioavailability and speciation can reduce this uncertainty and result in more accurate estimates of risk.

The bioavailability of Pb in soil is influenced by the species present, particle size, and whether Pb minerals have been encapsulated or coated by other mineral phases. For example, phases such as cerussite (PbCO3) and Pb oxide (PbO2) are more bioavailable than galena (PbS), anglesite (PbSO4), pyromorphite (Pb5[PO4]3Cl), and other encapsulated Pb phases (Zhang and Ryan 1999; Basta et al. 2001; Hettiarrachchi et al. 2001; Ruby 2004). Cerussite is a secondary Pb mineral commonly found in soils, but it is not stable and dissolves under acidic pH conditions. For these reasons, cerussite can be an important Pb mineral affecting Pb mobility and bioavailability in soils (Zhang and Ryan 1999). Lead associated with larger soil particles is generally less bioavailable than when attached to small particles, and encapsulation of Pb mineral phases with clays or quartz, for example, can also reduce Pb bioavailability (Chaney et al. 1989; Davis et al. 1993; Wixson and Davies 1993; Ruby et al. 1999).

The Cr species ingested affects its bioavailability. Chromium occurs primarily in 2 forms in soil: Trivalent Cr(III) and the more toxic hexavalent Cr(VI) (Bartlett and Kimble 1976; Langard and Norseth 1979; Bartlett 1991; Outridge and Scheuhammer 1993). The hexavalent form is more readily absorbed than the trivalent form, with Cr(III) having a bioavailability ranging from less than 0.5% to approximately 3% versus 10% for Cr(VI) based on orally administered doses in mammalian test species (IPCS 1988; De Flora and Wetterhahn 1989; O'Flaherty 1996). Hexavalent Cr is sometimes overestimated in soils, as the pH of soil extracts is adjusted with the oxidizing acid, HNO3, which has the potential to oxidize Cr(III) to Cr(VI) (James et al. 1995).

This investigation was performed to evaluate how the relative bioavailability of Pb and speciation of Pb and Cr from soils might affect assessment of risks to terrestrial receptors from exposure to these metals. The source of Pb and Cr posing these risks are soils remaining on a portion of the 184-acre former Col-Tex refinery site in Colorado City, Texas, USA, where an oil refinery operated from 1922 to 1969. This investigation focused on 2 areas, the Former Cracking Unit and Former Refinery. The Former Cracking Unit was where high-molecular-weight petroleum streams were broken down into lower-molecular-weight streams, and the Former Refinery area was where the initial distillation of oil and mixing of final shipped product (including Pb addition) occurred. Concentrations of Pb and Cr in soil were highest in the Former Refinery area (Figure 1). The most important and well understood cause of risk to terrestrial receptors is from exposure to contaminants by ingestion of soil attached to food or prey or through incidental ingestion of soil during grooming or preening of feathers or fur (USEPA 2005).

The screening-level ERA conducted at the site determined that Pb and Cr were the primary contributors to ecological risk estimates, possibly because the default assumptions concerning bioavailability and speciation overestimated exposure at the site. It was further determined that a substantial refinement of exposure and effects parameters, including bioavailability and speciation, was considered, and future soil conditions at the site were likely not to change. Taking these factors into consideration, the objective of this investigation was to determine the relative bioavailability of Pb and speciation of Pb and Cr in soils and to then reassess risks to the white-footed mouse (Peromyscus eremicus), eastern cottontail (Sylvilagus floridanus), American robin (Turdus migratorius), and northern bobwhite (Colinus virginianus). These receptors yielded the highest hazard quotients in a screening-level ERA at the site in which conservative assumptions regarding bioavailability and toxicity were made for these and several other receptors. It was intended that these new data would provide information needed to reduce uncertainty and increase accuracy in assessing risks from exposure to Pb and Cr in site soils. Reassessing risks would also provide new information on which to base more informed decisions about risk management at the site.

The objective of this study was to perform bioavailability and speciation analyses of Pb and Cr in soils as part of an ERA and determine the resultant effects on cleanup goals at this arid site. Our purpose in this manuscript is not to focus on the risk calculations per se but rather to elucidate the resultant change in cleanup concentrations and how they affected risk management when bioavailability adjustments are included. The bioavailability analysis included an extraction test that has been used to assess metals bioavailability in human health risk assessments but to our knowledge has not been similarly applied to assess risks to ecological receptors. As such, the applicability of using this approach and the associated uncertainty is discussed along with its potential application for assessing risk to other ecological receptors at other sites.


  1. Top of page
  2. Abstract
  7. Acknowledgements

Soil sampling, preparation, and characterization

The screening-level ERA identified 6 locations at the site where potential risks to ecological receptors from Pb in soils could not be ruled out (Fairbrother 2003). Similarly, 3 locations were identified where concentrations of Cr in soils might also pose risk. Ten soil samples were collected from these locations for analysis of Pb and Cr speciation and analysis of Pb bioavailability. Average measured soil parameters at the site included total organic carbon (2.1%), pH (7.6), and porosity (28.4%). The average particle-size composition for soil at the site was 8.4% clay/colloids (<0.005 mm), 30.6% silt (0.005–0.075 mm), 61% medium and fine sand (0.075–2.00 mm), and <0.1% gravel and coarse sand (>2.00 mm). The background concentration of Pb and Cr in nearby unimpacted surface soil was 6.7−1 dry weight and 7.8−1 dry weight, respectively.

Soil samples were collected from 0 to 15 cm in depth using a precleaned stainless-steel trowel or shovel. USEPA Method 3060A/7196 (USEPA 2003b; was used for Cr(VI) analysis. It consisted of extraction using alkaline digestion followed by colorimetric analysis. Total Pb was measured in both the bulk soil and the <250-μm fraction of the soil. The <250-μm soil fraction was used in the bioavailability analysis, as this size fraction is most likely to adhere to hands in humans (Duggin and Inkslip 1985), which was the rationale in developing the initial in vitro studies for human health risk assessments (Ruby et al. 1996). This smaller size fraction is also likely to adhere to fur and feathers, which would then be ingested by animals during grooming (Schoof 2003b). The result of these tests is a leachability of Pb, referred to as bioaccessibility. Bioaccessibility is a term used to describe the fractional dissolution of a metal from soil in an in vitro study. Measures of bioaccessibility are used to estimate relative bioavailability (Schoof 2003a).

Soil samples were submitted to Severn Trent Laboratories in Corpus Christi, Texas, for analysis of total Cr and total Pb using USEPA Method 3050B/6010B (USEPA 2003a;, which consisted of extraction by acid digestion followed by inductively coupled plasma-atomic emission spectrometry. Samples were also analyzed for organic Pb by Columbia Analytical Services in Kelso, Washington, USA, using the California LUFT method. This method is essentially a modified EPA 200.13 (USEPA 1997b) atomic absorption method for Pb on a xylene/MIBK extract of soil. Organic Pb is operationally defined as Pb removed from soil in a xylene extraction. This Pb will include Pb in methylated and ethylated forms, Pb acetate, and humically bound Pb.

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Figure Figure 1.. Soil sampling locations at the former Col-Tex refinery site, Colorado City, Texas, USA.

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In vitro procedure

The 10 samples were analyzed using the in vitro protocol developed and verified against animal models (Davis et al. 1992; Ruby et al. 1996). Each of the soil samples was oven-dried for 24 h and then sieved to <250 μm. A split of each sieved sample was analyzed for initial concentrations of Pb (Table 1). The remaining sample was used in the in vitro and quantitative electron microprobe analysis (EMPA).

Synthetic gastric solution was prepared by adding 530 μL 6 N hydrochloric acid, 1.25 g pepsin, 0.5 g citrate, 0.5 gmalate, 420 μL lactic acid, and 500 μL acetic acid per liter of deionized water, with the resulting solution having a pH of 2.5. Forty milliliters of gastric solution were combined in a 250-mL polyethylene separatory funnel with 0.40 g of each soil. The slurries were continuously purged with argon to simulate gastric mixing and maintained at 37 °C, a representative GI tract temperature in humans (Ruby et al. 1993). The mass of test material, volume of test solution, and pH of GI tracts (average during fasting and during ingestion of food) were based on examinations of stomachs and small intestines of rabbits (Ruby et al. 1993).

Subsamples of the slurry were collected after 20, 40, and 60 min, which were based on food transit times through the rabbit stomach (Ruby et al. 1993). At each sample interval, 2 mL of solution were removed and replaced with an equal volume of fresh gastric solution. Each subsample was centrifuged at 2,100 rpm (1,750 g) for 25 min to remove suspended particles >0.45 μm in diameter, and the supernatant was analyzed for Pb.

Microprobe procedure

To determine the soil mineralogy and solubility controlling phases of Pb, 10 samples were analyzed using EMPA at the Laboratory for Geological Studies, University of Colorado, Boulder, Colorado, USA, on a JEOL 8600 electron microprobe following the protocol of Davis et al. (1992). Briefly, polished sample pucks were prepared for EMPA by embedding 4 g of sample in epoxy within a sample mold, setting the molds to cure at room temperature, and grinding a flat surface on the sample side to expose as much sample as possible. All polishing steps used kerosene to avoid dissolution of water-soluble Pb phases and were performed at low speeds to avoid plucking of the sample grains. Finally, sample pucks were cleaned in an ultrasonic cleaner with isopropyl alcohol, air-dried, and placed in a carbon coater, where a thin layer of carbon is sputtered onto the surface of each puck.

Table Table 1.. Analytical results for lead (Pb) in soils (mg.kg1 wet wt)
AreaSampleTotal PbOrganic PbInorganic PbaInorganic Pb (%)Total Pb in Soil <250 μmBioavailability (%)
  1. a Inorganic Pb was calculated by subtracting organic Pb from total Pb.

  2. b Constituents not detected are shown as < reporting limit.

  3. c Mean values were calculated using one-half the detection limit for samples in which Pb was not detected.

Former cracking unitFC131<0 25b30.9>99.23728.2
Former refineryFR158,3004.658,29599.992,41077.8

Quantitative mineralogy was determined using wavelength dispersive spectrometers and mineral standards, and corrected using Phi Rho Z parameters. The Pb-bearing particles were identified using energy dispersive detection, wavelength dispersive detection, and backscatter electron image detection.


  1. Top of page
  2. Abstract
  7. Acknowledgements


Total concentrations of Pb in bulk soil in the 4 Former Cracking Unit samples ranged from 31 to 158−1 wet weight, with a mean of 92.8−1 (Table 1). Concentrations of organic Pb in these samples ranged from <0.25 to 1.0−1, indicating that the majority was inorganic Pb. On average, inorganic Pb in Former Cracking Unit samples was 99.4%. Concentrations of total Pb in the <250-μm soil fraction of Former Cracking Unit soil samples were greater than in bulk soil and ranged from 34.9 to 277−1, with a mean of 130−1.

Concentrations of total Pb measured in bulk soil samples from the 6 Former Refinery locations ranged from 105 to 58,300−1 wet weight (Table 1). Two of the 6 samples contained 35,100 and 58,300−1 Pb, while the remaining 4 samples contained between 105 and 238−1 Pb. However, concentrations in the <250-μm soil fraction of soil samples FR1 and FR2 were much lower than the bulk soil measurements, averaging only 9% of the total Pb concentration. The difference in concentrations in bulk soil as compared to the <250-μm soil fraction is due to the forms of inorganic Pb present, which is discussed in more detail in the following section. Concentrations of organic Pb in bulk soil were low, ranging from <0.25 to 4.6−1. As in Former Cracking Unit soil samples, inorganic Pb predominated in Former Refinery soils, averaging over 99% of the total Pb present.

In general, Pb, Cr, and other trace metals are usually associated with fine-grained particles in soils (Chaney et al. 1989; Horowitz 1991; Ruby et al. 1999), indicated by solid metal concentrations being generally higher in finer-sized fraction materials than in the bulk soil. In Former Cracking Unit samples, this same trend was observed. In samples FR1 and FR2, the concentrations of Pb in <250-μm fractions Pb is an order of magnitude lower than the bulk sample concentration (Table 1), likely because of the cementation of the samples by Pb carbonate minerals.

Inorganic Lead speciation

Two phases dominate the Pb mineralogy in the site soils: cerussite (Pb carbonate) and Pb-bearing iron oxide minerals (Table 2). Lead carbonate is the dominant phase in samples FR1 and FR2. This Pb likely is either from spillage of basic Pb carbonate or Pb oxide feedstock for tetraethyl Pb formulation or a weathering product of organic Pb compounds in soils. Most of this cerussite binds soil clasts together as cement. Given the high concentration of Pb in these samples and their nodular nature, this binding cerussite cement may account for the significantly greater concentrations of Pb in bulk samples relative to the <250-μm samples. In addition to cerussite, several samples contain Pb-rich calcite that is likely related to weathering of either organic or inorganic Pb in soil and subsequent calcite and/or caliche formation. Importantly, at neutral to slightly alkaline pH, Pb carbonate minerals are generally insoluble. However, at the acidic pH of the stomach, these minerals easily dissolve, and the Pb can be quite bioavailable. This high solubility under acidic conditions is the underlying reason for the high (>74%) bioavailability of Pb in samples FR1 and FR2 (Table 1).

Table Table 2.. Lead speciation in soil determined by electron microprobe analysis3
  1. a Blank cells indicate that no mineral phases were observed in point counts of that sample. FR = former refinery; FC = former cracking unit.

MineralRelative lead mass (%)
Clay10 1.66.5 0.3
Calcite11 7.5  5.7
Cerussite9295   45.2 
Fe oxide529980.388.551.087.2
Mn oxide11 7.4   
PbSiO4 1     
Fe sulfate0 11.5 1.36.8
Anglesite     2.5 
Lead phosphate    0.1  
Brass   1.0   
Solder   1.3   
Particles counted1682566046534762

The other major phases present in most samples were Pb-bearing iron oxides (Figure 2). Lead sorbs strongly to iron oxide compounds in soils (Dzombak and Morel 1990; Martinez-Villegas et al. 2004), so that weathered Pb from soluble phases is redistributed into iron oxide minerals in soils. In the 2 Former Cracking Unit samples analyzed (FC2 and FC4) and in 3 of the Former Refinery samples analyzed (FR3, FR4, FR6), the iron oxide-bound Pb dominates (Table 2). This sorbed Pb is bound even under relatively low pH conditions. Therefore, the lower observed bioavailability of these samples is consistent with the Pb speciation data.

Several minor Pb phases were also observed in soil samples (Table 2). These include Pb phases that are likely products of weathering (Pb bound to clay, Pb silicates, Fe-Pb sulfates, Pb phosphate) and industrial products containing Pb (brass and solder). The abundance of all these minerals is small enough that their contributions to total bioavailability are expected to be minor.

Lead bioavailability

Average results of in vitro testing showed that the 2 samples with the highest Pb concentrations had the highest Pb bioavailability, with the average bioaccessibility of these samples being 77% (Table 1). The samples with the highest average bioaccessibility were also the samples that had the highest cerussite content (FR1 and FR2). The average bioaccessibility of all other samples was <42% with bioaccessibility ranging down to 8%. The duplicate in vitro test results were within 13% of each other for all time periods. Matrix spike results indicated that spike recoveries for all samples were within 80%.


Concentrations of total Cr in the 4 Former Cracking Unit samples ranged from 18 to 494−1 wet weight, with a mean of 256−1 (Table 3). Concentrations of Cr(VI) ranged from <2 to 9−1 with a mean of 5.3−1. The concentrations of Cr(III) were calculated by subtracting the Cr(VI) from the total Cr concentration, yielding a range of 15 to 485−1 and a mean of 250.5−1; Cr(III) predominated in Former Cracking Unit samples, averaging 94.2% of the total Cr present.

Concentrations of total Cr ranged from 75 to 351−1 wet weight in the 3 Former Refinery samples, with a mean of 171−1 (Table 3). Concentrations of Cr(VI) in these same samples ranged from 4 to 8−1 with a mean of 6.3−1. The concentrations of Cr(III) were calculated as above and ranged from 71 to 343−1, with a mean of 165−1; Cr(III) prevailed in all samples, averaging 94.4% of the total Cr measured in Former Refinery soils. This result was not unexpected, as once Cr contacts soil, it is reduced to Cr(III), which is stable, insoluble, and immobile relative to Cr(VI) in soil conditions above pH 5 (Bartlett 1991; Losi et al. 1994).

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Figure Figure 2.. Photomicrograph showing cerussite and iron-lead oxide minerals in sample FR3.

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Table Table 3.. Chromium (Cr) analytical results in soils (units in mg.kg1 wet wt)
AreaSample IDTotal CrCr(VI)Cr(III)a% Cr(VI)% Cr(III)
  1. a Trivalent Lr (LrIII) was calculated by subtracting hexavalent Lr (LrVI) from total Cr.

  2. b Constituents not detected are shown as < reporting limit.

  3. c Mean values were calculated using one-half the detection limit for samples in which Cr was not detected.

Former cracking unitFC133<2b323.097.0
Former refineryFR4754715.394.4


The extraction test we used to estimate relative bioavailability of Pb and Cr to the ecological receptors for the site was originally developed for estimating relative bioavailability of metals to humans. Extraction test parameters such as extraction time, pH, and temperature can affect bioavailability estimates (Ruby et al. 1993, 1996). For this investigation, simulated gastric fluids were maintained at 3 7 °C, a representative GI tract temperature in humans (Ruby et al. 1993), which is similar to mammals including 7 Peromyscus species (36.5 °C), gerbil (Meriones unquiculatus; 36.9 °C), tree shrew (Tupaia belangeri; 36–39 °C), hamster (Mesocricetus auratus; 35.8 °C), 2 rat species (Ratus ratus; R. norvegicus; 36.7–38.1 °C), 2 ground squirrel species (Spermophilus tridecemlineatus; S. richardsonii; 34.1–37.5 °C), ferret (Mustela putorius; 38 °C), fox (Alopex lagopus; 38 °C), porcupine (Erethizon dorsatum; 38 °C), and raccoon (Procyon lotor; 38 °C; McNab and Morrison 1963; Frappell et al. 1992; Swenson and Reece 1993; Aiello and Mays 1998; Refinetti 1999; Yoda et al. 2000). This temperature (37 °C) is at the lower end of the body temperature range for birds, which ranges from about 37 to 42 °C (McNab 1966; Wasser 1986). While 37 °C is considered a representative body temperature for humans, it is known that actual body temperatures for other mammals and for birds will vary on the basis of several factors, including individual metabolism rate and heat loss rate. On a mass-to-mass basis, birds have higher body temperatures than mammals because they have higher rates of metabolism and usually have lower rates of heat loss than mammals (McNab 1966). For example, the body temperature reported for the American robin is 43.2 °C (McNab 1966). The difference in body temperatures between mammals and birds is a source of uncertainty in the use of these results to evaluate bioavailability in the mammalian and avian receptors at the site. This source of uncertainty can be addressed by increasing the gastric fluid temperature to about 40 °C to represent bird body temperatures. Because increases in the temperature of the simulated gastric fluid increases the bioaccessibility measured, the bioaccessibility values reported here for robin may be low. A general rule of thumb is that kinetically controlled reaction rates double with each 10 °C increase in temperatures (Espenson 1981). Therefore, a 3 °C difference from the test conditions should only increase bioavailability by less than about 30%.

The mass of test material (0.4 g) and volume of test solution (40 mL) were based on examinations of stomachs and small intestines of rabbits (Ruby et al. 1993). Extraction times of 20, 40, and 60 min coincide with the emptying time of rabbit stomachs (Ruby et al. 1993). In adult humans, the GI tract is 80% empty after 60 min following ingestion of a meal, and a child's stomach empties in 54 to 68 min (Ruby et al. 1996).

The pH value of 2.5 used to estimate GI tract pH is the average during fasting (pH = 1.0–1.3) and fed conditions (pH = 2.8–4.1) in rabbit stomachs (Ruby et al. 1993). These values are in general agreement with the GI tract pH of the rat, which has a stomach fluid pH of 1.0 to 1.5 under fasting conditions but otherwise ranges from 2.6 to 5.1 (Freeman et al. 1992). The pH in the forestomach of rodents often ranges from 4.0 to 4.5 (Karasov and Hume 1997). Rats usually nibble intermittently but continuously during the day, and when food is ingested, it causes the stomach fluid pH to rise because of the buffering capacity of the food (Wixson and Davies 1993). Similarly, stomach pH in pigs increased to 5.1, 4.0, and 3.1 at 30, 60, and 90 min after ingesting food (Wixson and Davies 1993). In ruminants such as cattle, stomach pH usually ranges from 3.5 to 4.5 (Chaney et al. 1989). The gastric pH of 5 species of falconiforms on a mouse diet averaged 1.7 (range −1.3–1.8) 4 h before meals and averaged 2.7 (range −1.5–3.5) 2 h after meals (Duke et al. 1975). In the same study, the gastric pH for 2 owl species 4 h before mouse meals was 2.35, ranging from 2.2 to 2.5, and was 2.75 (range −2.7–2.8) 2 h after feeding. Duke et al. (1975) also reported gastric pH data for the turkey (Meleagris gallopavo) and domesticated hybrid duck (Anas platyrhynchos × Cairina moschata), which averaged 3.0 premeal and 2.3 postmeal and 2.1 premeal and 2.1 postmeal, respectively. The pH used (2.5) is similar to or lower than that observed in the species of concern and in other species where gastric pH data are available, especially under nonfasting conditions. Therefore, pH conditions for the species studied resulted in a conservative estimate of pH since metals generally are more bioavailable at lower pH values. Thus, the method likely resulted in an overestimation of bioavailability for many species, thereby providing a higher risk estimate.

The GI system in birds shares with mammals the same fundamental components for digestion of food: A tubular intestine with proximal and/or distal fermentation chambers (Vispo and Karasov 1996; Karasov and Hume 1997). Minimal food (e.g., starch and fiber) digestion has been reported in the crops of some birds, with pH being as low as 4.5 (Ziswiler and Farner 1972; Vispo and Karasov 1996). The pH of gastric fluid secreted by the proventriculus has been reported to range from 0.2 to 1.2, while the pH in the ventriculus (or gizzard) ranges from 0.7 to 2.8 (Ziswiler and Farner 1972).

The amount of time that food travels through the digestive system from mouth to anus is termed the mean retention time. A summary of mean retention times for various species of birds indicates times ranging from approximately 45 to 90 min for nectar-, fruit-, or insect-eating birds; from approximately 60 to 190 min for leaf- and twig-eating birds; and from approximately 170 to 390 min for seed-eating birds (Karasov 1990; Levey and Karasov 1992, 1994; Karasov and McWilliams 2005). For the American robin, which is a seasonal frugivore, there is a difference in gut retention times depending on the food type. The mean retention time for fruit in the GI tract of the robin was 48 min, while for crickets it was 65 min (Levey and Karasov 1992). Although stomach emptying times or intestinal transit times were not reported, the actual times for these processes are expected to be less than the mean retention times.

Mean retention times in mammals are longer than in birds, with a general increasing trend in retention time with increasing body mass of the species. Mean retention times summarized by Stevens and Hume (1998) range from 3.4 h (vole) to 48 h (pig), with the actual retention time in the stomach to be less than this amount. For example, the 3 h required to completely empty a rabbit stomach (Ruby et al. 1993) is considerably less than the mean retention time of 27 to 39 h reported for rabbits for particles and fluids, respectively (Stevens and Hume 1998). The rabbit gut retention time of about 3 h is longer than retention times for other species, so bioavailability was likely overestimated. However, Ruby et al. (1993) calibrated their in vitro study to the rabbit model and subsequently found that these results were similar to later swine- and primate-based studies (e.g., Ruby et al. 1996).

Larger particle sizes would result in a smaller surface-to-mass ratio. As the rate of mineral dissolution is proportional to the particle size, the small particle size results in enhanced Pb dissolution (Davis et al. 1993). Larger particles would therefore dissolve less Pb. Since Pb concentrations were generally higher in the <250-μm fraction than in the bulk soil, the inclusion of data only from the <250-μm soil fraction in the extraction test likely increased Pb dissolution estimates in our study.

These data suggest that the assumptions used in the extraction test provide a reasonably accurate predictor of relative bioavailability and generate a reasonable estimate for the eastern cottontail while providing a conservative estimate for the other 3 species of concern because of the conservative body temperature, pH, and mean gut retention time values. Regulatory approval was granted by the Texas Commission on Environmental Quality (TCEQ), which considered this approach appropriate for assessing Pb and Cr risks at the site.

Until further research is done, however, extrapolation of in vitro bioavailability determinations to other species should be done with caution. While the current procedure mimics mammalian body temperatures of about 3 7 °C, it is less than avian body temperatures of 38 to 42 °C. Differences in gastric pH need to be examined for both bird and mammal species. The gastric pH of 2.5 may need to be lowered to account for gastric pH in some species of birds, such as falconiforms, and under fasting conditions. The sampling time of up to 60 min may not be sufficient to mimic gastric emptying time for some bird species. In ruminants, dietary intake 1st passes through the reducing environment of bacterial fermentation in the rumen, where complexation of metals can occur, changing the speciation and potential absorption in the upper GI tract.

Risk management

The data obtained from the in vitro and speciation studies were used to refine risk assessment calculations for site receptors and also used as input into risk management. As such, protective concentration levels (PCLs) were calculated for site soils according to the TCEQ ecological risk assessment guidance (TCEQ 2001). Based on the premise that a hazard quotient (HQ) of 1.0 is protective of a receptor exposed to a specific chemical, comparative PCLs were calculated. This PCL calculation method is based on the site-specific exposure dose and is a ratio of the target risk (in this case “1”) and the calculated risk (i.e., the HQ) multiplied by the chemical concentration. In this case, we used the 95% upper confidence limit (UCL) on the mean concentration. The 95% UCL soil concentration was divided by the HQ to yield the comparative PCL as follows:

  • equation image((1))

Likewise, PCLs based on the no-observable-adverse-effect level (NOAEL) and the lowest-observable-adverse-effect level (LOAEL; from Finley et al. 1976; Heinz and Haseltine 1981; Osborn et al. 1983; Zakrzewska 1988; Elbetieha and Al-Hamood 1997) were calculated from NOAEL- and LOAEL-based HQs as follows:

  • equation image((2))

In these PCL calculations, the chemical concentration represented by the estimated dose was attributed to the soil because soil was the abiotic medium contributing most of the dose for terrestrial receptors at the site. For example, soil had 16% and food items (i.e., vegetation and insects) 84% of the total estimated ingested Pb for the American robin (data not shown). Soil concentrations and wildlife toxicity thresholds used in calculating HQs were on a wet-weight basis. The arithmetic average of the NOAEL- and LOAEL-PCL for each receptor equals the comparative PCL for that receptor. The lowest of the comparative PCLs for each chemical becomes the final ecological PCL for that chemical (Table 4).

The final ecological PCL for both Pb and Cr was indicative of the soil concentration that should be protective of the most sensitive receptor (i.e., the most sensitive of the robin, quail, mouse, or rabbit). Thus, PCLs can be used as cleanup levels protective of the most sensitive ecological receptor exposed to soils.

Table Table 4.. Protective concentration levels (PCLs) used to manage lead (Pb) and chromium (Cr) risk to ecological receptors. All units in mg.kg1 wet weight
ReceptorComparative PCL for Cr(VI)Comparative PCL for Cr(III)Comparative PCL at 100% Pb bioavailabilityComparative PCL at 23.9% Pb bioavailability
American robin79.911251.578.3
Northern bobwhite3962,328160217
White-footed mouse58.610,3341,8202,135
Eastern cottontail22138,9975,3025,948

The data obtained from the extraction study were used to generate the PCL cleanup levels for Pb. Cleanup levels generated using site-specific bioavailability and speciation data were compared to cleanup levels based on the assumption of 100% bioavailability (Table 4). Considerable increases in Pb cleanup levels were achieved for all receptors, as the Pb PCL increased from 51.5 to 78.3−1. Similar results were obtained when site-specific speciation data for Cr were used. The comparative PCL for Cr nearly doubled from 58.6 to 112−1. For Pb, the change in bioavailability from 100% to 23.9% was the only contributor to the PCL change and resulted in a relatively consistent PCL increase across receptors examined. For Cr, however, the primary factor influencing the PCL change was toxicity reference values (TRVs; data not shown). The TRVs were originally based on studies where Cr(VI) species, such as potassium dichromate, were used; these were replaced with studies using Cr(III) species, such as chromium chloride. For mammals, TRVs based on studies using Cr(III) species were hundreds of times greater than TRVs based on Cr(VI) species; bird TRVs based on studies using Cr(III) were within 6-fold of TRVs derived using Cr(VI) species. This accounts for the nonlinearity of the increases in PCL values. For this same reason, the receptor for which estimated risks was greatest for Cr changed from the white-footed mouse to the American robin (Table 4).

These data provided evidence that site-specific bioavailability and speciation need to be considered when performing ecological risk assessments on soils contaminated with Pb and Cr. The costs associated with conducting the bioavailability and speciation studies were de minimus relative to the reduction in remediation costs realized at the site. The resulting PCLs were used to manage and segregate soils, allowing for more cost-effective ecological risk management at the site. While the 1.5- to 2-fold increase in Pb and Cr PCLs achieved here resulted in substantial reductions in soil remediation costs at the site, we realize that given the conditions and limitations of the data, achieving a less than 2-fold increase in PCLs may be less meaningful at other sites where Pb and Cr are of concern in soils. Whether such increases in PCLs are worth pursuing at other sites depends on site-specific conditions and risk management goals.


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  2. Abstract
  7. Acknowledgements

The results of this investigation indicated that site-specific measures of metals bioavailability and speciation should be used as part of assessing ecological risk to terrestrial receptors. Based on site-specific bioavailability of 23.9% for Pb, the calculated PCL for Pb was approximately 52% higher than was calculated using the conservative default bioavailability value of 100%. Similarly, the PCL for Cr based on site-specific speciation data almost doubled than when using the conservative default assumption that 100% of the Cr was Cr(VI). The reduced bioavailability of Pb and speciation of Cr and Pb resulted in increased protective cleanup concentrations for these metals in soils that in turn substantially decreased cleanup costs at the site.


  1. Top of page
  2. Abstract
  7. Acknowledgements

TOTAL PETROCHEMICALS USA, Inc., supported this investigation. The authors would like to thank John Slocomb and Todd Bridges for reviewing and providing constructive comments on the manuscript.


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  2. Abstract
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