Reversible Sorption and Irreversible Binding of Naphthalene and Î±

Environ. Sci. Technol. 1996, 30, 1205-1211

Reversible Sorption and Irreversible Binding of Naphthalene and r-Naphthol to Soil: Elucidation of Processes W I L L I A M D . B U R G O S , * ,† JOHN T. NOVAK,‡ AND DUANE F. BERRY§ Department of Civil and Environmental Engineering, The Pennsylvania State University, University Park, Pennsylvania 16802, and Department of Civil Engineering and Department of Crop and Soil Environmental Sciences, Virginia Polytechnic Institute & State University, Blacksburg, Virginia 24061

The sorption and irreversible binding of naphthalene and R-naphthol to two sandy soils were evaluated. To determine the extent that biologically-mediated oxidative coupling contributed to the irreversible binding of R-naphthol, the presence/absence of O2 and the presence/absence of biological activity were varied during the sorption period to operationally define biological coupling conditions. After a 2-d sorption period, sorbed material was removed by successive water and solvent extractions, alkali extractions, and combustion. Nonextracted material was defined as the amount of test compound remaining after solvent extraction (QNONEX). Naphthalene that was not extracted after the 2-d sorption period was caused by mass transfer-limited entrapment within micropores and was used to estimate the fraction of entrapped R-naphthol. The principle of superposition was applied to the R-naphthol results to determine the contribution of individual irreversible binding processes. Biologically-mediated oxidative coupling accounted for 49.4 ( 15.0% of QNONEX of R-naphthol. Oxidative coupling catalyzed by mineral surfaces accounted for 40.5 ( 15.8% of QNONEX of R-naphthol. Over contact times up to 200 d, nonextracted naphthalene increased from 1.6% to 11.7% of the initially sorbed material, coincident with the biological production of a more polar metabolite.

Introduction The irreversible binding of phenolic compounds to humic substances and activated carbon is well documented (14). The addition of a hydroxyl group to the aromatic ring * Corresponding author telephone (814) 863-0578, fax: (814) 8637304, e-mail address: [email protected]. † The Pennsylvania State University. † Department of Civil Engineering. § Department of Crop and Soil Environmental Sciences.

0013-936X/96/0930-1205$12.00/0

 1996 American Chemical Society

makes these compounds more reactive with mineral surfaces and soil organic matter (SOM) (5). The oxidative coupling of phenolics to SOM may account for their irreversible binding to soil. These covalent binding reactions can be catalyzed by clay minerals (6) and metal oxide surfaces (7) or biologically mediated by extracellular soil enzymes (8). During the aerobic degradation of polycyclic aromatic hydrocarbons (PAHs), hydroxylated intermediates are produced (9), and many aromatic metabolites are readily transported across cell membranes (10). The extracellular accumulation of PAH metabolites in culture media has been observed with naphthalene (11), phenanthrene (12), and six other PAHs (13). Therefore, these intermediates may also be oxidatively coupled to SOM or undergo further biodegradation. Stone (14) has proposed a mechanism for the adsorption and oxidation of phenolic compounds by manganese(III/ IV) oxide particles. Specific adsorption of the phenolic directly to the oxide surface is first required, followed by electron transfer, and then product release. Rates of reductive dissolution are primarily controlled by the adsorption of the phenolic and its willingness to lose an electron in the subsequent reaction (14). Reductive dissolution rates tend to increase with the addition of electrondonating substituents (e.g., -OH) and to decrease with the addition of electron-withdrawing groups (e.g., -Cl). Reaction rates also increase with decreasing pH and reach a maximum below the pKa of the phenolic compound (15). The reactive oxidized organic may be a free radical or a phenoxenium ion (16), but either species can form a covalent linkage to soil humic substances through an oxidative coupling reaction. Biologically-mediated oxidative coupling of phenolic compounds is an important synthesis route for the formation of humic substances in soil and lignin in woody plants. The two primary classes of enzymes involved in biological coupling reactions are peroxidase and phenoloxidase (i.e., laccase). Peroxidases have an absolute requirement for H2O2 for activity while phenoloxidases require O2 (8). Rates of oxidative coupling of phenols with peroxidase increase with the addition of electron-donating groups, which increase the potential number of reaction centers and/or provide increased resonance stability of the free radical (17). Suflita and Bollag (18) incubated R-naphthol with a fungal peroxidase and identified polynaphthols that were linked through C-C and C-O bonds. If R-naphthol is incubated with humic substances, we believe cross-coupled products would be formed in the same manner. Physical mass transfer limitations, such as retarded pore diffusion (19) or intraorganic matter diffusion (20), can produce apparent irreversible sorption. Mass transferlimited transport processes are distinct from covalent bond formation or chemisorption because no covalent linkage or charge transfer complex has been formed between the contaminant and the sorbent. If extremely long desorption periods are allowed, it would be expected that this physically-entrapped material would slowly gain entry into the bulk solution phase (21). Mass transfer-limited transport has also been used to explain the common two-stage desorption process observed in many sorption studies (22). Hydrophobic organic contaminant (HOC) molecules sorbed

VOL. 30, NO. 4, 1996 / ENVIRONMENTAL SCIENCE & TECHNOLOGY

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TABLE 1

Physical Characterizations of the Two Soils Studied sample location textural class particle size analysis % sand % silt % clay organic carbon contenta pHb surface areac (m2/g) pore volumec (mL/g) av pore radiusc (Å) % oxide surface contentd (mol of oxide/mol of surface)

high organic soil

low organic soil

beech/oak forest A horizon sand

cropped peanut field C1 horizon sand

89.9 8.9 1.2 3.20 3.80 1.11 0.0029 52.6 5.0

86.8 11.3 1.9 0.52 4.80 1.84 0.0064 69.0 5.2

a Determined by loss-on-ignition at 430 °C. b Measured with glass electrode in 1:2 (g of soil/mL of distilled water) mixture. c Measured by five-point N2 BET analysis in 0.05 relative pressure increments from P ) 0.10 to 0.30 Po. d Qualitative measure with scanning electron microscope performed with an hnu energy dispersive X-ray spectrometer system attached to a Camece Series 2 SEM.

to accessible sorption sites are readily desorbed, while HOCs held in diffusion-limited sites are released more slowly. The fate of organic contaminants released into soils or the subsurface may be affected by irreversible binding processes. Although PAHs are not readily susceptible to oxidative coupling, reactive metabolites are produced during the biodegradation of these compounds. Once the partial oxidation of a PAH occurs, oxidative coupling may become a competitive process with biodegradation. The objectives of this research were to (1) measure the extent of reversible sorption and irreversible binding of naphthalene and R-naphthol; (2) attempt to quantify the contribution of individual processes involved in irreversible binding; and (3) determine if the amount of soil organic matter affects irreversible binding processes.

Experimental Design In an effort to determine to what extent biologicallymediated oxidative coupling contributed to the irreversible binding of R-naphthol, adsorption conditions were varied to isolate this process: oxic-biotic experiments were conducted using non-autoclaved soil in an ambient (nonO2 limited) atmosphere; oxic-autoclaved experiments were conducted using autoclaved soil in an ambient atmosphere; anoxic-biotic experiments were conducted using nonautoclaved soil in an O2-limited, nitrogen atmosphere. Soils were autoclaved three times over 5 d for 30 min at 121 °C and 20 psi prior to use in the oxic-autoclaved experiments. Oxygen-free nitrogen, produced by passing N2 through an OxyTrap (Alltech) column, was used to purge all solutions and the contents of a Plexiglass glovebox. The anoxicbiotic experiments were set up and stored within the O2limited atmosphere of the glovebox. Because O2 and the appropriate extracellular enzymes are required to facilitate biologically-mediated oxidative coupling reactions, these reactions should not have occurred within the oxicautoclaved or anoxic-biotic systems.

Methods and Materials Materials. Radiolabeled naphthalene (1-14C) and R-naphthol (1-14C) purchased from Sigma Chemical with specific activities of 8.9 and 7.1 mCi mmol-1, respectively, were dissolved in ethanol and used without further purification. Unlabeled stock solutions for each test compound were prepared in 10 mM CaCl2 and filtered through 0.45-µm

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ENVIRONMENTAL SCIENCE & TECHNOLOGY / VOL. 30, NO. 4, 1996

FIGURE 1. Schematic of water and solvent extraction, and solvent extraction experimental procedures.

filters prior to use, and dissolved concentrations were verified by absorbance at 280 nm. A 10 mM CaCl2 solution prepared in distilled-deionized (d-d) water and buffered to pH 5.0 was used as the synthetic groundwater. The buffer contained 50 mL of 0.1 M potassium phthalate and 22.6 mL of 0.1 M NaOH/100 mL of d-d water and was added at 5% v/v to the CaCl2 solution. Two soils (thermic, coated, Typic Quartzipsamments) collected from Surry County, VA, were selected in order to minimize the effect of clay minerals on adsorption (Table 1). These two soils differed primarily in their organic carbon content (fOC) and are referred to in this paper as the high and low organic soils. The high organic soil (3.2% fOC) was collected from a beech/oak forest 0-15 cm below ground surface (bgs), and the low organic soil (0.52% fOC) was collected 30 m away from an adjacent cropped peanut field 15-30 cm bgs. The moist soils were passed through a U.S. standard sieve no. 10 and stored at 4 °C. Adsorption Experiments. Solvent Extraction Procedure. Three sets of triplicate batch soil reactors were used for each concentration of each test compound analyzed. Initial aqueous concentrations (Ci) of 10, 100, and 1000 µM R-naphthol and 1, 10, and 100 µM naphthalene were used. A schematic of the experimental steps and procedures employed is shown in Figure 1. Glass 13 × 100 mm screwcapped culture tubes with Teflon septa were prepared with 5.0 g of soil (dry wt), completely filled (ca. 7 mL) with a

solution of 14C-labeled and unlabeled test compound and synthetic groundwater to minimize any headspace within the reactor, and stored on a shaker table at room temperature for 2 d. Reactors were centrifuged, and 1-mL supernatant samples were collected and transferred directly into scintillation cocktail. The radioactivity within all samples was measured by liquid scintillation counting (LSC) (Beckman LSC 3510T; Beckman 14C standards were used to determine counting efficiency using the external standard method). The remaining supernatant volume was used for thin-layer chromatography (TLC) analysis using procedures detailed by Cerniglia et al. (23). One triplicate set was sacrificed at this time and immediately combusted within a 14C oxidizer (R. J. Harvey OX500 operated with O2 and N2 flow rates of 350 mL min-1 at a catalyst temperature of 710 °C and a combustion temperature of 925 °C for a 4-min combustion cycle). Aqueous controls with no soil were used to measure any physical losses within the reactors. The other two triplicate sets underwent a successive solvent extraction procedure to remove weakly held, physisorbed test compounds. Each reactor was filled with 4 mL of ethyl acetate (EtAc), and after 24 h the reactors were centrifuged and extract volumes were transferred directly into scintillation cocktail. This solvent extraction procedure was performed a total of five times. One triplicate set was sacrificed after the fifth solvent extraction and combusted within the 14C-labeled oxidizer. The remaining triplicate set underwent an alkali (0.5 N NaOH) extraction to remove test compounds associated with humic substances. Each reactor was filled with 4 mL of the aqueous alkali solution, and after 12 h the reactors were centrifuged, the supernatant was removed, and a second 4 mL of alkali was added. After the 24-h alkali extraction period, the reactors were centrifuged, the supernatant was removed, and the soil was combusted. Triplicate sets were sacrificed after each experimental procedure in order to estimate test compound recoveries throughout the experiments. This redundancy allowed high recoveries to be maintained for these volatile compounds: R-naphthol recoveries were 91100% and 87-100% for the high and low organic soils, respectively; naphthalene recoveries were 73-88% and 6281% for these two soils, respectively. The measured activity removed with the supernatant(s) was added to the activity measured upon sample combustion to calculate the total activity recovered. Aqueous controls revealed that adsorption to glassware was minimal and losses were primarily through volatilization during the various extractions. Water and Solvent Extraction Procedure. Another set of sorption experiments measured the extent of water extraction. Triplicate sets of soil reactors underwent successive water extractions (50 water rinses). After the first supernatant volume was removed and sampled, synthetic groundwater was added to the soil pellet and the reactor was remixed. The second supernatant volume was removed after 24 h and analyzed as previously described. After this point, the decanted supernatant volumes were pooled and analyzed after every fifth or tenth water extraction. During this stage of the desorption process, up to 10 water extractions were performed a day, and the water extraction procedure was completed over 7 d. After the final water extraction, these triplicate sets underwent the solvent extraction, alkali extraction, and combustion procedures outlined above.

Results and Discussion Naphthalene should not participate in biologically-mediated or surface-catalyzed oxidative coupling reactions in the test soils over the 2-d sorption period. While lignin peroxidases associated with the white rot fungus Phanerochaete chyrsosporium can oxidize PAHs (24), the ability of extracellular soil peroxidases to oxidize unsubstituted PAHs under environmental conditions is unproven. Additionally, phenoloxidases cannot utilize naphthalene as a substrate. The incubation of naphthalene with manganese oxide particles revealed that no chemical degradation of naphthalene occurred over a 9-week period (25). TLC analyses of the 2-d equilibrium supernatants revealed that no transformations of naphthalene had occurred during this time. Therefore, we contend that the nonextracted naphthalene in our 2-d experiments was due to mass transfer-limited processes. A number of terms need to be defined for the following discussion. The reversible component of sorption is operationally defined as the amount of sorbed material removed with the successive water and/or solvent extractions (QREV ) µmol/kg of soil). Nonextracted material is defined as the amount of material remaining after the final solvent extraction (QNONEX). Trapped material is defined as the fraction of material in micropores that is not extracted due to rate-limited processes (QTRAPPED). Bound material is defined as the remaining fraction of nonextracted material that is covalently bound to the soil (QBOUND). These terms are related by the following equations:

Qeq ) QREV + QNONEX

(1)

QNONEX ) QTRAPPED + QBOUND

(2)

and

where Qeq is the 2-d equilibrated sorbed concentration. Sorption Isotherms. Adsorption isotherm coefficients determined for R-naphthol and naphthalene with the two test soils are summarized in Table 2. Estimates of log KOC values based on correlations with the compound’s octanolwater coefficient and solubility (26) range from 1.87 to 2.08 for R-naphthol and from 2.60 to 2.74 for naphthalene and are in good agreement with the results obtained in this study. In order to compare the water extraction desorption isotherms for the different initial aqueous concentrations tested, the results have been normalized to the equilibrated sorbed and aqueous concentrations (Figure 2). The final data points on the y-axis correspond to the nonextracted concentrations (QNONEX) remaining after the fifth solvent extraction. The greatest amount of nonextracted R-naphthol (QNONEX/Qeq) 0.33-0.67 for the three concentrations) occurred under Oxic-Biotic conditions where biologicallymediated coupling reactions likely occurred. Under oxicbiotic conditions, the greater extent of irreversible binding at the lower R-naphthol concentrations may have been due to at least two factors. Surface saturation of the available irreversible binding sites or substrate saturation of the oxidative coupling enzymes may have occurred at the higher concentrations, allowing a greater percentage of material to be irreversibly bound at the lowest concentration. The results obtained under anoxic-biotic conditions (Figure 2b) are included to contrast desorption patterns for conditions where biological coupling did not occur.


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TABLE 2

Linear Adsorption Isotherm Coefficients

a

sorbate

soil

adsorption conditions

no. of data points

KD (L/kg)

r2

log KOC (L/kg of OC)

R-naphthol R-naphthol R-naphthol R-naphthol R-naphthol R-naphthol naphthalene naphthalene

high high high low low low high low

oxic-biotic anoxic-biotic oxic-autoclaved oxic-biotic anoxic-biotic oxic-autoclaved oxic-biotic oxic-biotic

11 6 6 8 4 4 12 9

7.73 (0.22)a 5.16 (0.42) 5.76 (0.17) 0.396 (0.02) 0.456 (0.01) 0.291 (0.01) 18.2 (0.59) 0.96 (0.06)

0.98 0.95 0.99 0.96 1.00 0.99 0.97 0.94

2.38 2.21 2.26 1.88 1.94 1.75 2.75 2.27

Standard error of regression coefficient given in parentheses.

TLC analyses revealed that 4.6 ( 2.7% (mean ( SD) of R-naphthol activity extracted from the high organic soil under oxic-biotic conditions was measured as a more polar transformation product, while the same procedure measured only 0.53 ( 0.22% of R-naphthol transformation products under anoxic-biotic or oxic-autoclaved conditions. A reduction in biological activity, as evidenced by decreased production of metabolites, may have reflected a general decrease in enzymatic activity, and we have used this response as a general indicator of extracellular oxidoreductase activity. If biologically-mediated oxidative coupling reactions were reduced under anoxic-biotic and oxic-autoclaved conditions, then we contend that mineral surface-catalyzed reactions accounted for the binding of R-naphthol under these conditions. Qualitative, high magnification SEM/ X-ray spectrometer analyses of these soils identified a number of iron and titanium oxides. Qualitative, elemental topology was determined with lower magnification and revealed that roughly 5% (molar basis) of the soil surfaces were coated with metal oxides (Table 1). Therefore, a catalytic amount of metal oxides may have been present in these soils to account for the observed irreversible binding under the anoxic-biotic and oxic-autoclaved conditions.

FIGURE 2. r-Naphthol and naphthalene water extraction desorption isotherms with the high organic soil. Sorbed (Q) and aqueous (C) concentrations obtained during desorption have been normalized to the equilibrated sorbed (Qeq) and aqueous (Ceq) concentrations. The final open symbols represent the sorbed concentrations remaining after the fifth solvent extraction (QNONEX).

Under anoxic-biotic and oxic-autoclaved conditions, the amount of nonextracted R-naphthol was reduced; however, a significant amount of irreversible binding still occurred (QNONEX/Qeq ) 0.14-0.41 for both conditions). Enzyme assays to verify the presence/absence of peroxidase and phenoloxidase under the different experimental conditions were unsuccessful, and thus involvement of oxidative coupling enzymes could not be verified. TLC analysis of the equilibrated supernatant was used to determine if any transformations of the test compound occurred. These

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Naphthalene desorption isotherms with the high organic soil are shown in Figure 2c. The water extraction was unable to remove a portion of the sorbed naphthalene however, solvent extraction was able to recover virtually all of the remaining material (QNONEX/Qeq) 0.016-0.024). Thus, naphthalene sorption to the high organic soil was nearly fully reversible and showed no concentration effect on the amount of bound material. The amount of naphthalene that remained with the soil after water and solvent extractions was probably associated with micropores and not extracted due to rate-limited mass transfer processes. Distribution of Sorbed Species. Results obtained with R-naphthol from experiments using the solvent extraction procedure are presented in Table 3 and Table 4. The amount of nonextracted material with the high organic soil was consistent with the water extraction desorption isotherms. The oxidative coupling mechanisms suggest the binding of the contaminant to SOM including humic substances. The activity recovered with the alkali extract revealed that a fraction of the irreversibly bound material was incorporated into the humic fraction. Common humic substance extractions use a 1:5 (g of soil/mL of alkali solution) ratio (27), while we used a 1:1.6 ratio to work within the culture tube reactors. The alkali extraction procedure used for this study may have reduced the humic substance extraction efficiency, which would lead to an

TABLE 3

Summary of Sorption Data for r-Naphthol with the High Organic Soil sorbed concentrationb remaining after adsorption conditions oxic-biotic oxic-biotic oxic-biotic anoxic-biotic anoxic-biotic anoxic-biotic oxic-autoclaved oxic-autoclaved oxic-autoclaved

2-d aqueous equilibriuma

2-d sorbed equilibriumc

solvent extractionc

alkali extractionc

153 ( 11 n ) 12 7.18 ( 0.39 n ) 15 0.47 ( 0.03 n ) 21 180 ( 10 n ) 12 10.4 ( 0.6 n ) 12 0.83 ( 0.05 n ) 12 186 ( 8.6 n ) 12 10.8 ( 0.5 n ) 15 0.84 ( 0.02 n ) 12

1170 ( 17 n)3 128 ( 6.9 n)6 13.1 ( 0.20 n)6 1130 ( 61 n)3 124 ( 6.3 n)3 12.6 ( 1.0 n)3 1110 ( 30 n)3 121 ( 5.2 n)6 12.6 ( 0.4 n)3

376 ( 4.0 n)6 69.4 ( 1.5 n)6 8.80 ( 0.58 n)9 184 ( 21 n)6 38.4 ( 1.9 n)6 5.19 ( 0.30 n)6 152 ( 6.8 n)6 38.3 ( 1.3 n)6 3.83 ( 0.21 n)6

307 ( 9.2 n)3 57.1 ( 5.6 n)3 8.21 ( 0.69 n)6 136 ( 6.4 n)3 28.7 ( 2.0 n)3 3.38 ( 0.26 n)3 130 ( 3.7 n)3 33.2 ( 0.1 n)3 3.12 ( 0.15 n)3

a Aqueous concentrations reported in µmol/L and represent mean ( SD for n observations determined by LSC of supernatant sample. concentrations reported in µmol/kg. c Mean ( SD for n observations determined directly by sample combustion.

b

Sorbed

TABLE 4

Summary of Sorption Data for r-Naphthol with the Low Organic Soil sorbed concentrationb remaining after adsorption conditions oxic-biotic oxic-biotic oxic-biotic anoxic-biotic anoxic-biotic anoxic-biotic oxic-autoclaved oxic-autoclaved oxic-autoclaved



solvent extractionc

alkali extractionc

794 ( 18 n)9 68.9 ( 9.2 n ) 12 5.49 ( 0.06 n ) 18 752 ( 21 n)9 76.4 ( 2.8 n ) 12 5.93 ( 0.25 n)9 827 ( 12 n)9 77.3 ( 1.8 n)9 7.59 ( 0.26 n)9

284 ( 2.2 n)3 42.9 ( 2.2 n)3 6.23 ( 0.24 n)6 342 ( 30 n)3 38.3 ( 1.4 n)3 5.61 ( 0.24 n)3 239 ( 15 n)3 31.4 ( 0.7 n)3 3.32 ( 0.03 n)3

100 ( 13 n)6 24.2 ( 2.1 n)6 4.44 ( 0.07 n)6 65.7 ( 6.4 n)3 13.1 ( 1.0 n)6 2.83 ( 0.03 n)3 33.8 ( 3.0 n)3 8.80 ( 1.4 n)3 0.78 ( 0.06 n)3

89.2 ( 2.7 n)3 20.0 ( 3.9 n)9 2.29 ( 0.40 n)6 65.3 ( 1.8 n)3 5.10 ( 0.30 n)3 1.38 ( 0.40 n)3 24.4 ( 4.5 n)3 6.70 ( 0.10 n)3 0.50 ( 0.06 n)3


underestimate of activity within this material, or a significant amount of binding to the nonextractable humin fraction occurred. However, these results do confirm that a portion of the bound material was directly associated with humic substances, presumably through an oxidative coupling reaction. Direct chemisorption of certain organics to mineral surfaces has been demonstrated; however, this was unlikely with R-naphthol. Yost et al. (28) used infrared spectroscopy to illustrate which the binding mechanism of salicylate to goethite was a chelate structure which involved the coordination of both the carboxylic and hydroxyl substituent groups. Because R-naphthol possesses only a single hydroxyl group, it cannot form a chelate structure with soil mineral surfaces. Charge transfer complexes have been proposed for the adsorption of phenolics to activated carbon

b

Sorbed

(3); however, because the pKa of R-naphthol is 9.3, there would be virtually none of the required phenolate anion for this to occur at pH 5 in these experiments. The general trends on the extent of irreversible binding were also seen with the low organic soil (Table 4). The greatest amount of bound R-naphthol occurred under oxicbiotic conditions, and a greater extent occurred at lower concentrations. Unlike the high organic soil, TLC analyses revealed that virtually no biotransformation of R-naphthol occurred under any condition. The limited biotransformation of R-naphthol suggests that the presence and activity of extracellular enzymes (including peroxidase and phenoloxidase) may have been limited in this soil, but because the greatest amount of bound material occurred under oxicbiotic conditions, biological activity still appeared to promote irreversible binding.


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TABLE 5

Summary of Sorption Data for Naphthalene under Oxic-Biotic Conditions sorbed concentrationb remaining after soil

sorption period

low

2d

low

2d

low

2d

high

2d

high

2d

high

2d

high

10 d

high

30 d

high

200 d



solvent extractionc

alkali extractionc

0.46 ( 0.02 n)9 4.23 ( 0.27 n)9 59.2 ( 2.50 n ) 12 0.09 ( 0.04 n)3 0.83 ( 0.07 n)9 7.13 ( 0.30 n ) 18 6.52 ( 0 n)9 6.78 ( 0 n)9 5.12 ( 0 n)9

0.75 ( 0.01 n)3 7.96 ( 0.49 n)3 56.4 ( 0.6 n)3 1.26 ( 0.04 n)3 12.7 ( 1.6 n)3 128 ( 10.5 n)6 129 ( 2.0 n)3 129 ( 4.6 n)3 131 ( 3.3 n)3

0.06 ( 0.02 n)3 0.66 ( 0.04 n)3 4.8 ( 0.4 n)6 0.03 ( 0.001 n)3 0.31 ( 0.03 n)3 2.1 ( 0.5 n)6 2.9 ( 0.1 n)3 7.8 ( 0.3 n)3 15.4 ( 0.1 n)3

0.04 ( 0.01 n)3 0.57 ( 0.07 n)3 3.3 ( 0.1 n)3 0.02 ( 0.001 n)3 0.24 ( 0.07 n)3 1.8 ( 0.2 n)6 2.5 ( 0.5 n)3 5.4 ( 1.6 n)3 3.9 ( 0.1 n)3


Results obtained with naphthalene from experiments using the solvent extraction procedure are shown in Table 5. Virtually all of the naphthalene sorbed to the high organic soil during the 2-d period was recovered upon solvent extraction. The remaining sorbed material (1.6-2.4% of Qeq) was believed to be due to rate-limited diffusive processes. For the low organic soil, the percentage of nonextracted naphthalene increased to 7.6-8.3% of Qeq. Physically the low organic soil had a greater unit surface area, larger interior pore volume, and less organic matter coverage compared to the other soil to provide more access to interstitial sites. Superposition of Adsorptive Processes. The principle of superposition allows the effects of combined processes to be determined separately and then added together to determine the combined result. This principle can only be applied to processes that are linearly related and assumes the results of one process does not affect the conditions for the other processes. Based on our experimental design, we have applied the principle of superposition to differentiate the contribution of individual processes to the overall irreversible binding of R-naphthol. For the following discussion, we will assume that all the operative sorption processes can be described by linear isotherms. Based on the results obtained with naphthalene, roughly 8% (8.13 ( 0.32) of the material sorbed to the low organic soil and 2% (2.04 ( 0.38) of the material sorbed to the high organic soil was trapped within mass transfer-limited sites and not covalently bound. Using naphthalene as a measure of the extent of rate-limited diffusive processes in these soils and assuming R-naphthol will exhibit similar diffusive characteristics, these percentages of sorbed material can be subtracted from the nonextracted fraction to estimate the remaining percentage that was bound according to eq 2. With the high organic soil, QTRAPPED for R-naphthol was estimated to equal 0.02Qeq, and with the low organic soil, QTRAPPED was estimated as 0.08Qeq. This distinction revealed that mass transfer-limited processes accounted for 4.3 ( 1.6% of QNONEX of R-naphthol sorbed under oxic-biotic conditions in the high organic soil and 16.1 ( 5.9% in the low organic soil for the three concentrations tested.

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b

Sorbed

Adsorption superposition was further used to estimate the contribution of biologically-mediated and mineral surface-catalyzed oxidative coupling to the overall irreversible binding of R-naphthol. General comparisons can be made between the different experimental conditions to evaluate the extent of each process. Under anoxic-biotic and oxic-autoclaved conditions, we assumed that the operative irreversible binding process was surface-catalyzed, while under oxic-biotic conditions, we assumed that both processes could occur. Therefore, the difference between QBOUND calculated for the oxic-biotic condition and the QBOUND values for the anoxic-biotic and oxicautoclaved conditions can be attributed to biologicallymediated oxidative coupling by the following relationship:

QBOUND ) QBIO + QMINERAL

(3)

where QBIO is the covalently bound concentration due to biological coupling, and QMINERAL is the covalently bound concentration due to surface-catalyzed oxidative coupling reactions. Biologically-mediated coupling of R-naphthol was responsible for 49.0 ( 7.3% of QNONEX sorbed under oxicbiotic conditions in the high organic soil and for 49.7 ( 21.0% of QNONEX in the low organic soil. The standard deviation of this estimate was based on results obtained for the three concentrations tested under the anoxic-biotic and oxic-autoclaved conditions (n ) 6). Mineral surfacecatalyzed oxidative coupling was responsible for 46.7 ( 8.0% of QNONEX in the high organic soil, and 34.2 ( 19.9% of QNONEX in the low organic soil. Mineral surfaces are commonly assumed to be covered by SOM when fOC exceeds a few percent (29), yet our results show that mineral surfacecatalyzed reactions were actually more important in the high organic soil. This suggests that the associations of SOM with clay minerals and metal oxides may not necessarily cover inorganic surfaces, but may promote chemically-induced oxidative coupling due to the proximity of the required reactive soil constituents. Environmental Significance. The results obtained with R-naphthol suggest that PAH metabolites are quite sus-

94% of the aqueous activity was naphthalene at any time tested; and (2) naphthalene recoveries were always greater than 83% for all contact times, suggesting that little 14CO2 was evolved and lost. Therefore, we believe these results demonstrate that irreversible binding may become competitive with biodegradation.

Literature Cited

FIGURE 3. Competitive pathways between further contaminant biodegradation and irreversible binding after the initial oxidation of a parent PAH.

ceptible to oxidative coupling and can occur in both organic and mineral soils. During the degradation of naphthalene and PAHs in general, extracellular metabolite accumulation has been observed (11-13). The irreversible binding of PAH metabolites may then become a competitive process with biodegradation (Figure 3). The impact of oxidative coupling on the fate of naphthalene was studied by comparing the extractability of this compound over extended adsorptive contact times. Reactors with naphthalene (Ci ) 100 µM) and the high organic soil were prepared under oxic-biotic conditions and allowed to equilibrate for 2, 10, 30, and 200 d. At the end of the contact time, the solvent extraction procedure was used to measure the extent of irreversible binding. Results of the naphthalene contact time experiments (Table 5) confirmed that, once the partial oxidation of a PAH occurs, the irreversible binding of metabolites can occur. After 2 d only 1.6% of Qeq was nonextracted. Between 10 and 30 d, the microbial community in this soil began to degrade naphthalene. TLC analyses of the 30and 200-d supernatants revealed the production of an unidentified, more polar metabolite (Rf ) 0.6). Coincident with this increase in naphthalene utilization, the nonextracted fraction increased to 6.0% of Qeq after 30 d and continued to increase to 11.7% of Qeq after 200 d. Identical reactors prepared with autoclaved soil (oxic-autoclaved) served as sterile controls up to 30 d and showed no increased binding. Over the 200-d contact time, we believe little mineralization of naphthalene occurred for two reasons: (1) although metabolite production occurred, no less than

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Received for review June 19, 1995. Revised manuscript received November 6, 1995. Accepted November 14, 1995.X ES950428B X

Abstract published in Advance ACS Abstracts, February 1, 1996.


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Reversible Sorption and Irreversible Binding of Naphthalene and Î±

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