The influence of sediment resuspension on the degradation of phenanthrene in flow-through microcosms

MARINE

Marine Environmental Research 61 (2006) 202–223

www.elsevier.com/locate/marenvrev

ENVIRONMENTAL

RESEARCH

The influence of sediment resuspension onthe degradation of phenanthrene in

flow-through microcosms

Lawrence A. LeBlanc *, Jeanne D. Gulnick 1,Bruce J. Brownawell, Gordon T. Taylor

Marine Sciences Research Center, State University of New York at Stony Brook,

Stony Brook, NY 11794-5000, United States

Received 17 November 2003; received in revised form 21 September 2005; accepted 10 October 2005

Abstract

The effect of sediment resuspension on the mineralization of phenanthrene was examined inmicrocosms and sediment slurries. In computer-controlled, flow-through microcosms, 14C-phenan-threne-amended sediments were resuspended into overlying oxic water at frequencies of 12, 4, 1,0.25 and 0 d�1. In slurry bottle experiments 14C-phenanthrene-amended sediments were continu-ously resuspended under oxic (excess air headspace) and anoxic (N2 headspace) conditions and min-eralization was measured at periods from 2 h to 7 days. Our main findings were: (1) mineralizationrate constants from the microcosms ranged from 0.001 to 0.01 d�1 and increased with frequency ofresuspension, (2) these rates fell between those measured in oxic and anoxic slurries and were pre-dicted within a factor of 2.5 by a model in which mineralization depended on the degree of oxygenexposure, and (3) the phenanthrene-degrading bacterial community was more active in resuspendedsediments incubated in the microcosms than in sediments which were not resuspended, or which werestored under refrigeration. We conclude from these experiments that the effects of sediment resuspen-sion on phenanthrene degradation are consistent with a primary role of average oxygen exposure,and also an alteration in the PAH-degrading activity of microbial populations.� 2005 Elsevier Ltd. All rights reserved.

0141-1136/$ - see front matter � 2005 Elsevier Ltd. All rights reserved.

doi:10.1016/j.marenvres.2005.10.001

* Corresponding author. Present address: School of Marine Sciences, 5741 Libby Hall, University of Maine,Orono, ME 04469-5741, United States. Tel.: +1 207 581 4376; fax: +1 207 581 4990.

E-mail address: [email protected] (L.A. LeBlanc).1 Present address: Saint Joseph�s College, 237 Whites Bridge Road, Standish, ME 04083, United States.

mailto:[email protected]

L.A. LeBlanc et al. / Marine Environmental Research 61 (2006) 202–223 203

Keywords: Biodegradation; Phenanthrene; Bioavailability; PAH; Sediment resuspension; Microcosm

1. Introduction

The effect of sediment resuspension on the remobilization and bacterial mineralization ofhydrophobic organic contaminants (HOCs) in the environment has received little attention.Resuspension of marine sediments is common in harbor areas that experience disturbancefrom tidal currents, storms, shipping, boating, and dredging activities and may represent asignificant mechanism for the loss of HOCs from contaminated harbors. Polycyclic aro-matic hydrocarbons (PAHs) are a class of non-ionic hydrophobic compounds that enteraqueous environments through oil spills, point and nonpoint discharges, and atmosphericdeposition (Neff, 1979). Once in the water column, they sorb onto particles that settle andbecome part of the sediments; those bound to soot particles may be especially persistent(Gustafsson, Haghseta, Chan, Macfarlane, & Gschwend, 1997; McGroddy, Farrington,& Gschwend, 1995). PAHs are of environmental concern because of their persistence andbecause some compounds and their metabolic byproducts are toxic and/or carcinogenic(Nordqvist et al., 1980). Mechanistic studies which attempt to elucidate the primary con-trols limiting loss of PAH from sediments are needed in designing more effective manage-ment or remediation strategies for contaminated harbors and estuaries.

Sediment resuspension is expected to have a number of effects on PAH mineralizationby bacteria. By exposing suspended sediment to an oxic water column and increasing thedepth of oxygen penetration into highly reducing sediment beds, resuspension can enhanceaerobic respiration, the primary means of bacterial PAH degradation (Cerniglia & Heitk-amp, 1989). The dissolved concentration of hydrophobic chemicals generally controls therate of microbial degradation, and therefore both the extent and rate of desorption ofsorbed compounds can be important rate-limiting processes to bacterial transformation(Guerin and Boyd, 1992; Harms and Zehnder, 1995; Knaebel, Federle, McAvoy, & Vestal,1994, 1996; Mihelcic and Luthy, 1990; Ogram et al., 1985). Sediment resuspension hasbeen shown in natural systems to increase the flux of sediment-sorbed pollutants to theoverlying water column (Latimer, Davis, & Keith, 1999; Millward, Morris, & Tappin,1998; Sanders, Taylor, & Jones, 1996). The mixing of sediment particles due to reuspen-sion can lead to changes in grain size distribution, precipitated mineral phases and organiccoatings (Alkhatib & Castor, 2000; Simpson, Apte, & Batley, 1998), which can affect par-titioning into interstitial waters. Finally, physical and biological disturbance of sedimentshas been shown to stimulate the physiological activity of heterotrophic bacterial popula-tions in both laboratory and field studies (Findlay, Trexler, Guckert, & White, 1990; Find-lay, Trexler, & White, 1990) and may also affect PAH-degrading, bacterial populations.

The present study examines the effect of sediment resuspension on the degradation rateof 14C-labeled phenanthrene in flow-through microcosms. Phenanthrene was used as amodel PAH because of its widespread distribution in the environment, being producedfrom both petrogenic and pyrogenic sources (LaFlamme & Hites, 1978) and because ofits intermediate hydrophobicity (Schwarzenbach, Gschwend, & Imboden, 1993). Increas-ing the frequency of sediment resuspension in an experimental system would be expectedto increase sediment oxygen exposure and rates of mineralization. Therefore it washypothesized that the rate of phenanthrene mineralization would increase as a function

204 L.A. LeBlanc et al. / Marine Environmental Research 61 (2006) 202–223

of increased sediment resuspension due to increased exposure of the sedimentary bacteriato oxygen in interstitial and overlying waters. To test this hypothesis, experiments wereconducted in flow-through microcosms and sealed slurry bottles. In the microcosms,organic-rich, highly-reducing sediments were resuspended over a range of frequencies intoan overlying oxygenated water column. Batch slurry experiments were conducted underboth oxic and anoxic conditions to better understand the role of intermittent oxygen expo-sure of bedded sediments on phenanthrene degradation in the microcosms. Microcosmresults were modeled as a linear combination of oxic and anoxic rates determined fromthe slurry experiments, based upon estimates of oxygen exposure at different frequenciesof resuspension in the microcosms.

2. Materials and methods

2.1. Materials

The 9-14C-phenanthrene with a specific activity of 3.07 · 104 Bq/mmol was purchasedfrom Sigma Chemical Company, St. Louis, MO. Solvents were Fisher certified, pesticidegrade (Fisher Scientific, Pittsburgh, PA) and Burdick and Jackson, glass distilled, pesticidegrade (Scientific Products, Edison, NJ). PAH standards for gas chromatographic analysiswere purchased from Ultra Scientific, North Kingstown, RI.

2.2. Sediment collection and preparation

Sediment was collected from a depositional area adjacent to a sand and gravel offload-ing dock in Port Jefferson Harbor, Long Island, NY. Port Jefferson Harbor receiveshydrocarbon inputs from an oil-fired power plant as well as from commercial (ferry andbarge traffic) and recreational boating activities. Previous work demonstrated that sedi-ments from nearby sites contained bacterial communities acclimated to degrading PAHs(Kerr & Capone, 1988). The top 10 cm sections from five cores (8 cm in diameter) werecombined and passed through a 500 lm mesh sieve to remove macrofauna. The sediment,which was black, fine-grained, muddy sand, was allowed to settle for 1 week under refrig-eration (�4 �C) prior to use in the microcosms.

2.3. Experimental flow-through system

A flow-through, sediment–water microcosm system was developed that allowed for thecontrolled resuspension of sediment into an overlying water column (Fig. 1). The systemconsisted of a 55 gallon plastic seawater reservoir, a multichannel pump, a flow distribu-tion panel, 15 1 L glass microcosms, magnetic stir motors, a water bath, oxygen electrodeand PC controller.

During an experiment, sediments were quiescent and the PC controller and distributionpanel directed seawater flow through the microcosms at a rate of 1–2 mL min�1 (residencetimes = 0.3–0.7 d). During programmed mixing events flow was suspended. After 3 min ofmixing, sediments settled for 30 min under no-flow conditions tominimize loss of fines, afterwhich the inflow was restored by the PC controller. Chamber effluent was directed by sole-noid to either 4 L amber glass sample bottles, or to an oxygen probe to measure and recorddissolved oxygen concentrations. Oxygen measurements (computer-programmed) were

Fig. 1. Schematic of the flow-through microcosm system. (a) Overall view of the system, with tubing connectingmicrocosms and solenoid valves (depicted as circles and crosses, respectively) shown for the three microcosms onthe right, and tubing connecting solenoid valves and the plenum shown for three microcosms on the left, (b)diagram of a single microcosm showing inflow tubing (made of Teflon, denoted as TFE) and outflow tubing(made of stainless steel), sample port, oxic, and anoxic sediment layers and magnetic stir bar overlain by filteredseawater. Inflow is halted during sediment resuspension. Outflow can be directed either to waste, an oxygen probeor to sample collection bottles (see text for details).


used to confirm that overlying waters never became anoxic. Time-dependent drift in the O2

probe was corrected with periodic measurements of the air-saturated inflow seawater. Thefrequency and duration of mixing, settling, flow redirection and O2measurements were pro-grammed prior to the initiation of the experiment. All changes of state, e.g. mixing, settling,flow conditions, and O2 electrode potentials were automatically recorded by the PCcontroller.

2.4. Effects of sediment resuspension in flow-through microcosms

An experiment was conducted to test the effects of different frequencies of resuspensionon the initial mineralization rates of radiolabeled phenanthrene in the flow-through micro-cosms. 14C-phenanthrene (40 lL at 0.48 Bq lL�1) dissolved in ethanol was added to eachempty microcosm chamber for a total activity of 18.5 Bq and a total mass of 107 lg 14C-phenanthrene per chamber. After the ethanol evaporated, sediment was added to eachmicrocosm (34 g dry weight) along with 100 mL of 1 lm-filtered seawater. Microcosmswere maintained at 0–1 �C in an ice-water bath to minimize bacterial activity and stirredperiodically (stirred for 3 min, every 12 min) under computer control for 48 h. At the end


of the equilibration period, the microcosms were completely filled with filtered seawater.To initiate the experiment, the ice was removed and the bath provided with continuouslyrunning tap water (18–20 �C). An equilibration time of 48 h was chosen to minimize theaging effect observed with longer contact times and to maximize bioavailability.

Sediments in triplicate microcosms were suspended at frequencies of 0, 0.25, 1, 4, and 12d�1. Water samples were collected in amber glass bottles from each chamber outflow for 8 hevery 2 days and subsampled to quantify mineralization to 14CO2. To each sample collec-tion bottle, 2 N KOH (10 mL) and mercuric chloride (1%) were added prior to collection toprevent the loss of 14CO2 through volatilization and to prevent further degradation of 14C-phenanthrene or metabolites in the outflow. Water samples were also extracted to deter-mine 14C-phenanthrene concentration in whole water (dissolved plus suspended solids).

Sediments were sampled during the course of the experiment via a glass tube insertedthrough the sampling port. These samples were centrifuged in 10 mL glass tubes, thesupernatant water discarded, and the sediment frozen until ready for analysis.

2.5. Sediment slurry bottle incubations

In order to estimate the upper and lower limits of mineralization, slurry incubations wereperformed with 14C-phenanthrene and Port Jefferson sediment with either a large air head-space (oxic conditions) or with N2 gas in the headspace (anoxic conditions). Mineralizationwas determined by sacrificing bottles at several time-points over a 2 week period, and mea-suring the evolved 14CO2 via serum bottle respirometry (described in Section 2.7).

In addition to the incubations described above, slurry incubations were also performedwith 14C-phenanthrene and unlabeled Port Jefferson sediment that had been aged in flowthrough microcosms for 1 month. This was done in order to determine whether resuspen-sion and/or conditioning in the flow-through microcosms affected the resident bacterialpopulation in terms of their ability to degrade a representative PAH (i.e. phenanthrene).Sediments were conditioned at 18–20 �C for 30 days under two conditions: never disturbedand frequently resuspended (12 d�1). Incubations were performed over a short (2 d) timeperiod, and compared to slurry incubations with the same sediment which had not been inthe flow-through microcosms but stored at 4 �C over the same time period. Incubationsbegan approximately one hour after the addition of the 14C-phenanthrene substrate, inorder to minimize sequestration and maximize bioavailability for the purpose of charac-terizing resident, phenanthrene-degrading bacterial populations.

2.6. Analysis of background PAH and total organic carbon

Field sediments were analyzed for background PAH concentration via gas chromatog-raphy (GC) and gas chromatography/mass spectrometry (GC/MS). For purposes ofdescribing the total PAH loading of field sediments, the sum of 16 unsubstituted PAHcompounds from the EPA priority pollutant list and commonly available as a standardmixture was referred to as total PAH. The extraction procedure was a modification ofthe method described in Lamoureux, Brownawell, and Bothner (1996). Sediments wereextracted three times with 30 mL of a 1:1 mixture of hexane:acetone, using a 600 W ultra-sonic processor (4710 series, Cole Parmer Instrument Co., Chicago). The extract was par-titioned against solvent-extracted, deionized water, reduced in volume, and charged ontoan open glass column containing 7 g of 5% deactivated silica gel (Biosil, 100–200 mesh,


Fisher Scientific). An aromatic fraction (40 mL of 20% dichloromethane in hexane) waseluted from column and reduced to approximately 1 mL for instrumental analysis. Anal-yses were performed on a Hewlett Packard 5890 GC and a Hewlett–Packard 5890A/5710GC/MS. The analytical column was a 30 m DB-5 (J + W Scientific, Folsom, CA, 0.2 lmfilm thickness, 0.32 mm ID), the carrier gas was nitrogen (flow rate 2 mL/min), and theoven ramping program was 100–250 �C at 10 �C/min. Para-terphenyl was added as a sur-rogate prior to extraction and ortho-terphenyl was added as a recovery standard immedi-ately prior to instrumental analysis. Sediment samples were also measured for organiccarbon content on a Carlo Erba model N1500 CHNS analyzer, after treatment with2 N HCl to remove carbonate-containing minerals.

2.7. Determination of mineralization in flow-through microcosms and slurry incubations

Mineralization of 14C-phenanthrene was determined in the flow-through microcosmsby measuring evolved 14CO2 in the outflow water via serum bottle respirometry. Samplesof water (75 mL) from each microcosm were placed in 150 mL serum bottles. Each bottlecontained a filter paper (Whatman no. 1) soaked with 0.5 mL of 2 N KOH to trap evolved14CO2 (base trap) and was sealed with a teflon-backed butyl rubber stopper and crimpingcap. 14CO2 and all other forms of inorganic carbon were driven into the headspace by add-ing 4 mL of 2 N HCl and agitating on a shaker table (150 rpm) for 60 min. Filter paperswere then removed from the bottles and placed in scintillation cocktail (Hionic Fluor,Packard Company, Meriden, CT) and counted on a Rackbeta Model 1217 liquid scintil-lation counter (LKB Wallac, Finland). Samples were corrected for quench using the exter-nal standard ratio method. Recoveries using this procedure were between 92 and 95% asverified using 14C-labeled, sodium bicarbonate in seawater.

Mineralization of 14C-phenanthrene in sediment slurry incubations was also measuredvia serum bottle respirometry. Sediment (2 g dry weight) and 1 lm-filtered seawater(10 mL) were added to a series of autoclaved 150 mL serum bottles. 14C-phenanthrene dis-solved in ethyl alcohol (50 lL of 0.04 Bq/lL) was added to each bottle via syringe, and thebottles were sealed as described above. Sample bottles were wrapped in aluminum foil toexclude light, and agitated on a shaker table (150 rpm) during the incubations. For theoxic experiments the large (150 mL) air headspace in the bottles provided oxic conditionsfor the duration of the experiments. Anoxic conditions for the anaerobic slurries were pro-duced by sparging seawater and sediment slurry with N2 gas and sealing the bottles underan N2 headspace. At selected times 4 mL of 2 N HCl was injected into the slurry and thebottles agitated on the shaker table for 60 min to terminate bacterial activity and liberate14CO2. Filter papers were removed and counted for radioactivity via the liquid scintillationcounter (LSC). Control samples consisting of sediment and seawater autoclaved twice for1 h at 260 �C were subjected to the same treatment.

2.8. Extraction of 14C label

Aqueous concentrations of 14C-phenanthrene and metabolites were determined by sol-vent extraction of subsamples from the microcosm effluent. At selected times (1–2 d), 600–1000 mL of outflow water was collected, reduced to a pH of 2 with 12 NHCl, and extracted(2 mL · 150 mL ethyl acetate, 2 mL · 150 mL hexane). Radioactivity of extract aliquotswas measured via LSC.


Concentrations of phenanthrene and metabolites were determined in microcosm sedi-ments by extracting sediments (0.2 g dry weight) sequentially with 2 mL · 2 mL ethyl ace-tate, 1 mL · 2.5 mL hexane, and 2 mL · 2.5 mL acetone. Extractions were performed inglass centrifuge tubes, using both vortexing and sonication (Bransonic model 5200 sonicbath, Branson Cleaning Equipment Company, Shelton, CT). Between each extraction,samples were centrifuged and the supernatant removed. Extracts were combined and analiquot counted via LSC.

Radiolabel remaining in the sediment after solvent extraction (residual fraction) wasdetermined by combustion. Extracted sediments were combusted at 900 �C using anOX-600 Biological Oxidizer (Harvey Instrument Corporation, Patterson, NJ) and theeffluent was trapped in scintillation cocktail and assayed for radioactivity via LSC.

Selected aqueous and sediment extracts were further analyzed to determine the relativeconcentration of transformed metabolites by thin layer chromatography (TLC, silica 60 Aparticle size, 250 lm layer thickness, Whatman International Ltd., Maidstone, England).Areas of the plate were scraped and counted via LSC to determine the relative concentra-tion of phenanthrene and more polar products. Plates were developed using 5% methylenechloride in hexane as the mobile phase. The location of non-metabolized phenanthrene onthe plate was verified using unlabeled compound and ultraviolet light detection on sepa-rate plates, and by adding unlabeled compound directly to the sample prior to spotting.

2.9. Statistical analysis

Analysis of covariance and Tukey�s multiple range test was conducted on mineraliza-tion data from the flow-through experiments using JMP Statistical Software (SAS Insti-tute, Carey, NC). Slopes of regression lines were compared statistically within eachexperiment using analysis of covariance with time as the covariate. Pairs of slopes weretested for significant differences using Tukey�s multiple range test (Zar, 1984). Differenceswere considered significant at P < 0.05.

2.10. Model of oxygen-mediated mineralization

To test the hypothesis that the primary effect of sediment resuspension was to increasephenanthrene mineralization through increased oxygen exposure, a simple first orderkinetic model of phenanthrene mineralization was constructed. This model depended onlyupon aerobic and anaerobic rates of phenanthrene mineralization determined indepen-dently from the slurry incubations described above, as well as estimates and measurementsof the duration of oxygen exposure and the depth of oxygen penetration into the sedimentsat the different resuspension frequencies. The model statement is as follows:

oCot

¼ ½fOzKO þ fIOzfIOtKO þ fIAzfIAtKA�C; ð1Þ

where:

oCot : the mineralization rate of 14C-phenanthreneðday�1Þ,fOz: the fraction of the sediment bed which is permanently oxic,fIOz: the fraction of the sediment bed which is intermittently oxic,fIAz: the fraction of the sediment bed which is intermittently anoxic,


fIOt: the fraction of time that fIOz is intermittently oxic,fIAt: the fraction of time that fIAz is intermittently anoxic,KO: the aerobic mineralization rate constant (determined from oxic slurries),KA: the anaerobic mineralization rate constant (determined from anoxic slurries), andC: concentration of 14C-phenanthrene added.

Eq. (1) assumes that there are only two intrinsic rates of degradation for phenanthrene(aerobic and anaerobic). Under this assumption, an apparent degradation rate can be pre-dicted by multiplying the intrinsic rates by the percent of the sediment bed (oxic and/oranoxic) and the fraction of time spent in either one or both those of those two states.

2.11. Estimates of oxygenation in resuspension experiment

In order to obtain estimates of the volumes of sediment which were oxic and anoxic(fOz, fIOz, and fIAz) and the amount of time that these portions of the bed remained in thesestates (fIOt and fIAt) at the different frequencies of resuspension, a number of measure-ments were made in small-scale experiments conducted with Port Jefferson sediment inthe flow through microcosms.

Sediment bed depth and the depth of the apparent redox boundary (sediment colorchange) and depth of oxygen penetration were measured during the flow-through experi-ment, as well as in separate trials. In static (flow-suspended) chambers containing 34 g dryweight of Port Jefferson sediment, measurements of bed depth and depth of the apparentredox boundary were made 1, 2, 3, 5, 6 and 12 h, 3 days and 7 days after a single resus-pension in which the overlying water was kept oxic with continuous aeration. In addition,the depth of oxygen penetration was estimated using an Eh probe (Microelectrodes Inc.,Londonderry, NH) in flow-through microcosm chambers after a single resuspension. Howquickly the redox boundary formed after a single resuspension event was also estimated.Several measurements of Eh were made in the chambers over a 2–3 day period.

3. Results

3.1. Background PAH concentrations

Blank concentrations were always a negligible percentage (�1%) of the total PAH con-centrations. The total concentration of unsubstituted PAHs in Port Jefferson Harbor sed-iments was 10.3 ± 0.3 lg g�1 (mean ± standard deviation, n = 5, collected on differentdates) indicative of an area with moderate to high loadings of PAH contamination(NOAA, 1985). The organic carbon content of this sediment was 3.70 ± 0.15% (n = 3).

In flow-through microcosm and slurry bottle experiments 14C-phenanthrene was notadded at trace concentrations. The amount of 14C-phenanthrene added to sediments inthe flow-through experiment (3.2 lg g�1) was 10 times the ambient phenanthrene con-centration (0.29 ± 0.27 lg/g) and a third of the total ambient PAH concentration. Addi-tion of 14C-phenanthrene was lower in the slurry bottle experiments (1.1 lg/g). Because14C-phenanthrene was added in excess concentrations, and given the much greaterdesorbability and bioavailability of the spike relative to the native compound (Carmi-chael, Christman, & Pfaender, 1997), the rates of 14C-phenanthrene mineralization werenot significantly affected by the natural phenanthrene levels. However, as discussed


below, it appeared that the microbial populations were well acclimated to PAHsubstrates.

3.2. Mineralization in flow-through microcosms

The mineralization rate of 14C-phenanthrene increased with resuspension frequency(Fig. 2(a)). Cumulative mineralization after 10 days in the two most frequently resus-pended treatments (4 and 12 d�1) was 3–4 times greater than in the least frequently resus-pended treatments (0 and 0.25 d�1; Fig. 2(a)). The absence of a lag period in the frequentlyresuspended treatments (1, 4 and 12 d�1) indicated the presence of well-acclimated PAH-degrading bacterial populations. Despite evidence for enhancement of phenanthrene min-eralization after 9–10 days in the two least frequently resuspended treatments, the data fitreasonably well to a first order kinetic model. Slopes of the resulting lines from the first 11days were taken to represent first order initial mineralization rate constants and had r2 val-ues ranging from 0.78 to 0.99 (Fig. 3). Rate constants were 5–10 times greater for the 4 and12 d�1 resuspension treatments when compared to the 0 and 0.25 d�1 treatments (Table 1).These slopes were compared using analysis of covariance with time as the covariable. Dif-ferences among mineralization rates were highly significant (P < 0.00001) for the first 11

14C

O2

(% o

f in

itial

)

0

5

10

15

20

25

Time (days)

0 5 10 15 20 25 30

Aqu

eous

14C

(%

of

inita

l)

0

2

4

6

8

10

a

b

12 d-1

4 d-1

1 d-1

0.25 d-1

0 d-1

Resuspension Frequency

Fig. 2. Cumulative loss of 14C from the sediments in the flow-through experiment (a) to mineralization (14CO2)and (b) to the aqueous phase (dissolved + particulates). Flow-through microcosms contained 34 g of sedimentlabeled with 14C-phenanthrene resuspended at various frequencies. Cumulative values in (a) were calculated bysumming values from triplicate microcosms and computing the mean and standard deviation for each time point.Because of less replication, cumulative values in (b) were calculated by summing values from 1 to 3 replicates ateach time point.

Time (Days)

0.0 2.5 5.0 7.5 10.0 12.5

Ln

[P] s

-0.08

-0.06

-0.04

-0.02

0.00 12 d-1

1 d-1

0 d-1

Least squares regression

Fig. 3. Mineralization data plotted as the natural logarithm of substrate disappearance over time for three of thefive resuspension frequencies from the flow-through experiment. Rates were derived from least squaresregressions of ln-transformed mineralization data, expressed as ln [P]S, where [P]S is the fraction of non-mineralized 14C-phenanthrene remaining at time t. The slope of each line represents the first order mineralizationrate for the first 11 days. For illustrative purposes only the 0, 1 and 12 d�1 resuspension frequencies are shown.

Table 1Comparison of phenanthrene degradation rates obtained from flow-through slurry experiments

Source Experimental design k (d�1) Turnover time (d)

Resuspension experiment (d�1)

0 Flow through 0.001 10000.25 Microcosm 0.002 5001 0.005 2004 0.01 10012 0.009 111

Anoxic slurry 0.007 140Oxic slurrya Slurries 0.075 1330 Day microcosm sediment 0 d�1 0.139 730 Day microcosm sediment 12 d�1 0.274 4Refrigerated sediment 0.072 14

Literature values

Heitkamp and Cerniglia (1987) Flow throughPristine ecosystem Microcosm 0.006 180PAH-impacted ecosystem 0.025 40

Shiaris (1989)a SlurriesWeymouth back river 0.127 8Fort point channel 0.051 20Lower neck 0.119 8

McElroy et al. (1994)Spectacle island Slurries 0.063 16Fort point channel 0.025 40Buzzard�s bay 0.056 18

a Averaged over all seasons.


days of all treatments. Results from the Tukey�s multiple range test between slope pairsshowed significant differences between the 12 d�1 and the 1, 0.25 and 0 d�1 treatmentsand between the 4 d�1 and the latter three treatments. Comparisons of 12 d�1 vs. 4 d�1

as well as 0 d�1 vs. 0.25 and 1 d�1 were not significantly different. Mineralization rates


were comparable in magnitude to published rates measured in sediments from hydrocar-bon-impacted areas (Table 1).

3.3. Mineralization in slurry incubations

Mineralization rates of 14C-phenanthrene measured in oxic and anoxic slurry bottleexperiments bracketed the rates measured in the flow through-microcosm studies (Table1). Mineralization proceeded rapidly with no lag period for the oxic slurries, suggestingthe presence of bacterial populations acclimated to aerobic PAH degradation. In contrast,a lag period in phenanthrene mineralization was observed for the anoxic slurries. Initialmineralization rates conformed reasonably well to a first order model (r2 = 0.98 and0.71 for the oxic and anoxic slurries, respectively). Rates were comparable to those mea-sured in studies of other sediments with high PAH concentrations (Table 1), such as thestations from Boston Harbor measured by Shiaris (1989) and McElroy, Shiaris, and Cap-uzzo (1994).

Phenanthrene-mineralizing activity was enhanced in slurry bottles containing sedimentsthat previously had been incubated in the flow-through resuspension chambers (Table 1).Sediments resuspended 12 times d�1 for 30 days mineralized 14C-phenanthrene in oxicslurries twice as fast as comparable undisturbed sediments. Both sets of incubated sedi-ments had faster mineralization rates (1.8 and 3.6 times faster for 0 and 12 d�1 treatments,respectively) than sediment stored at 4 �C and not incubated in the flow-through micro-cosms (Table 1).

3.4. Degree of oxygenation: short term flow-through and static microcosm experiments

The time to re-establish redox conditions following a resuspension event was deter-mined by measuring the depth of the redox boundary with an Eh probe following a dis-turbance. Suboxic conditions (measured as a negative Eh) began forming as soon as3 min after resuspension ceased, although a distinct sediment–water interface was notobserved until 30 min after cessation of resuspension. After 6 h the redox boundary fellto a depth between 0.5 and 2 mm for all microcosms. Based upon these observations,the time to re-establish a redox boundary was estimated to be 10 min over the first 10 daysof the experiment, which was the time over which the initial mineralization rates weredetermined. The intermittently oxic/anoxic portion of the sediment bed was assumed tobe sufficiently oxygenated for aerobic metabolic processes to occur for 13 min per resus-pension event (3 min during resuspension, plus 10 min of settling and re-establishmentof redox conditions). For the purposes of model calculations it was assumed that dynamicsof oxygen utilization in sediment was independent of resuspension frequency.

The measurements described above were used to model initial degradation rates as afunction of the degree of oxygenation of the sediment bed at each resuspension frequency.Model predictions, based on independent measurements of mineralization in oxic andanoxic slurry incubations and estimates of oxygen exposure, matched rates measured inthe flow-through microcosms at different frequencies of resuspension, within a factor of2.5 (Fig. 4). This result is consistent with the notion that oxygen is mediating the rate of min-eralization at different frequencies of sediment resuspension. The ability to predict the mag-nitude of degradation rates in the flow-through microcosms from rates determined in thebatch slurry experiments is an important result that will be discussed in the sections below.

0

0.0025

0.005

0.0075

0.01

0.0125

Min

eral

izat

ion

Rat

e (d

-1)

12 4 1 0.25 0

Resuspension Frequency (d-1)

Predicted Rate

Measured Rate

Fig. 4. Comparison of initial mineralization rate constants with model predictions based upon potential oxygenexposure for the flow-through resuspension experiment. See text for further explanation of model predictioncalculations.


3.5. Effects of resuspension on phenanthrene removal

Radioisotope inventories accounted for the majority of the 14C-phenanthrene spike(86% ± 30%) and mass balances were sufficient to allow identification of the major routesof loss of 14C-label from the system (Fig. 5(a)–(c)). The cumulative loss of 14C-phenan-threne to the overlying water was 8% or less of the total label for all treatments(Fig. 2(b)), less than lost to mineralization (up to 20% cumulative loss in 12 d�1 resuspen-sion treatment) but still an important pathway of loss in the system. Frequently suspendedsediments (4, 12 d�1 treatments) lost four times as much to the aqueous phase as less fre-quently resuspended sediments (0, 0.25 and 1 d�1 treatments). Concentration of the 14C-label in sediments declined over time and revealed an inverse relationship between solvent-extractable activity and resuspension frequency (Fig. 5(a)–(c)). For all resuspension fre-quencies, between 15% and 17% of the initial label was incorporated into a non-extract-able (residual) phase by day 30 (Fig. 5(c), Table 2). The incorporation of 14C into theresidual phase was of similar magnitude to the amounts converted to 14CO2. However,unlike mineralization, there was not a strong trend of increased incorporation to the resid-ual phase with increased resuspension frequency (see Fig. 6).

3.6. Fate of transformation products

To further characterize the fate of metabolites in the flow-through experiments, selectedextracts were analyzed for parent and metabolite compounds by TLC. For aqueousextracts in resuspended sediments, transformed products made up a significant percentageof the total label (23% ± 9%, n = 7) early to midway into the experiment (days 3–11) thatfell to low levels by the end of the experiment (4.7% ± 0.2%, n = 2). In undisturbed andlow frequency resuspension treatments (0 and 0.25 d�1) percent metabolites remainedlow throughout the experiment (4% ± 1%. n = 4).

The proportion of the total 14C-label recovered as metabolites increased from 3% (day 0)to 28% (day 11), declining to 3% (day 30) in the most frequently resuspended treatment

Fig. 5. Radiolabel inventory, expressed as the relative percentage of 14C-label present in cumulative CO2 andaqueous phases, as well as solvent-extractable (Solvent Extr) and non-extractable (Residual) phases in sedimentsafter (a) 0 days, (b) 11 days, and (c) 30 days.

Table 2Effects of time and resuspension frequency on the incorporation of 14C-label into metabolites vs. the residualphase of sediments in flow-through microcosms

Sampling day Fractiona Resuspension frequency

12 d�1 0 d�1

0 M 0.02 (3%) 0.01 (1.6%)0 R 0.02 (3%) 0.02 (3%)11 M 0.17 (28%) 0.10 (16%)11 R 0.08 (13%) 0.04 (7%)14 M 0.04 (7%) 0.05 (8%)14 R 0.06 (11%) 0.04 (7%)30 M 0.02 (3%) 0.05 (8%)30 R 0.10 (17%) 0.09 (15%)

Data are presented as nmoles phenanthrene and % of initial 14C-phenanthrene added (in parentheses).a M: solvent-extractable metabolites, R: non-extractable residual fraction.


Time (days)

0 5 10 15 20 25 30Solv

ent-

extr

acta

ble

activ

ity (

% o

f in

ital)

0

20

40

60

80

100 12 d-1

4 d-1

1 d-1

0.25 d-1

0 d-1

Resuspension Frequency

Fig. 6. Solvent-extractable activity from sediments over time, expressed as the percent of the initial activityadded. Values are the means from duplicate microcosms.

14CO2 (moles)

0.00 0.05 0.10 0.15

Res

idua

l Sed

imen

t Pha

se (

mol

es)

0.000

0.025

0.050

0.075

0.100

0.125y = 0.577x, r2 = 0.63

Fig. 7. Comparison of the concentration of 14C-label in the residual (solvent non-extractable) phase withcumulative 14CO2 concentrations for all resuspension treatments. The equation for the linear regression (leastsquares fit) and the r2 value are also presented.


(Table 2). In contrast, the proportion of the 14C-label in the residual (non-extractable) sed-iment fraction increased steadily from 3% (day 0) to 13% (day 11) to 17% (day 30). Similar,though less dramatic changes were observed in the 0 d�1 treatment. These results suggestthat metabolites disappeared over time andweremineralized and/or possibly converted intoresidual phase. Interestingly, the amount of 14C-phenanthrene converted to 14CO2 corre-lated well with the amount incorporated into the residual phase (r2 = 0.63, P < 0.01, Fig. 7).

4. Discussion

The results from the flow-through microcosm experiments showed conclusively thatsediment resuspension stimulated mineralization of 14C-phenanthrene. Higher frequencies


of sediment resuspension had faster rates of phenanthrene degradation, supporting thehypothesis that increased oxygenation was largely responsible for the enhanced phenan-threne degradation. Furthermore, evidence for the stimulation of PAH-degrading bacte-rial populations was provided by enhanced mineralization in aerobic slurry bottleassays with sediments frequently resuspended in the flow-through microcosms, when com-pared to microcosm sediments that were not resuspended or sediment that had been storedunder refrigeration. This can be expected to provide additional control on the observedrates, especially later in the time course of the experiment.

4.1. Effect of oxygen on phenanthrene degradation

Comparison of measured rates of 14C-phenanthrene to independent model calculationsare consistent with the hypothesis that oxygen exposure is an important control on PAHdegradation in our resuspension chambers. Estimates of oxic and anoxic mineralizationrate (KO and KA from Eq. (1)) were derived from slurry bottle experiments. The effectof suboxic conditions modifying oxic degradation rates was not considered. However,Bauer and Capone (1985) demonstrated that degradation rates at even low oxygen ten-sions (10% of saturation) were similar to rates at fully saturated conditions. Oxygen asa master controlling variable of PAH degradation has been discussed by several authors(DeLaune, Patrick, & Casselman, 1981; Freijer, de Jonge, Bouten, & Verstraten, 1996;Mille, Mulyono, El Jammal, & Bertrand, 1988) and oxygen concentration is an importantparameter in degradation models (e.g. Freijer, 1996).

This model predicted rates of phenanthrene mineralization reasonably well, beingwithin a factor of 2.5 of actual observations in the worst case (Fig. 4). Model calculationswere able to describe the effects of resuspension frequency on the magnitudes of degrada-tion rates. Batch slurry incubations provided reasonable estimates of the intrinsic degra-dation rates that occur in sediment beds of episodically disturbed, flow-through studies.The significance of this modeling exercise is not the mathematical model itself, which issimplistic. More sophisticated kinetic models of degradation take into account a numberof important environmental variables, including bacterial biomass and growth rate, rate ofoxygen delivery, and rate of dissolution and desorption of substrate (Borden & Bedient,1986; Guerin & Boyd, 1992; Ogram et al., 1985; Reible, Kalliat, & Thibodeaux, 1991;Subba-Rao & Alexander, 1982). What is significant is that independent measurementsof aerobic and anaerobic mineralization rates determined from slurry incubations couldbe used, along with estimates of oxygen exposures, to predict rates in bedded microcosmsystems. Furthermore, according to the model the effects of widely different resuspensionrates on phenanthrene mineralization are relatively modest (e.g. 4·).

One uncertainty in comparing mineralization rates determined in bedded sediments tothose determined in batch slurry incubations is the proper ratio of sediment to water to usein slurries to approximate porewaters in bedded sediments. In the sediment bed, the sed-iment: water ratio is higher than in the slurry bottle exposure and is partly a function ofporosity. We have found that phenanthrene mineralization increases with the sediment:water ratio, up to a value of 1:10, after which further increase yields no further increasein mineralization rate (LeBlanc, 2001). Both the slurry bottles and bedded sediments fromthese experiments had sediment: water ratios above 10. Thus the consistency in microbialmineralization rates between these different exposure conditions supports the use of theslurry bottle rates in our interpretation.


Although the effect of oxygen on phenanthrene mineralization was predictable, neitherpredicted nor measured rates varied by more than a factor of 10. This was probably due tothe limited oxygenation attainable on sediment particle surfaces during resuspension of thehighly reducing sediment, which had both stored chemical oxygen demand and a high bio-logical oxygen demand. This may also be the reason for the similarity seen between the 4d�1 and 12 d�1 resuspension treatments. Because of high oxygen demand in the water col-umn, it is possible that resuspending sediment at a greater frequency than 4 d�1 did notprovide any more oxygen at the particle surfaces (the site of degradation). Low oxygenconcentration, coupled with high oxygen demand in the sediment also caused the rapidre-establishment of redox conditions after the 3 min resuspension event. In spite of thehigh biological and chemical oxygen demand exerted on the water column by the resus-pended sediments, dissolved oxygen never fell below 20% saturation (1.6 mg L�1), andwas mostly 80–90% of the saturated standard (6.5–7.3 mg L�1) throughout theexperiment.

4.2. Phenanthrene sorption and bioavailability

It is generally believed that the sorbed fraction of PAH compounds is not available forbacterial uptake, and hence, desorption into the aqueous phase may be an important rate-limiting step to bacterial uptake and subsequent compound transformation (Cerniglia &Heitkamp, 1989; Mihelcic, Lueking, Mitzell, & Stapleton, 1993). Several studies suggestthat the extent and rate of desorption of hydrophobic contaminants from soils and sedi-ments may significantly limit rates of microbial degradation (Guerin and Boyd, 1992;Harms and Zehnder, 1995; Knaebel et al., 1994, 1996; Ogram et al., 1985). Efforts tomodel the fate (including transport and biodegradation) of hydrocarbons in porous mediahave either assumed local equilibria between particles and interstitial waters (Chen, 1993;Reible et al., 1991) or that desorption is rapid compared to the transport of oxygen (Bor-den & Bedient, 1986).

To test the assumption of local equilibrium (between sediment and porewater) a first-order estimate of the characteristic time for pore water equilibration due to desorptionwas derived using the model of Wu and Gschwend (1986). Recent studies have shown thatthis model, in some cases, does not provide reasonable predictions of contaminant desorp-tion from field-aged sediments, and is incompatible with resistant desorption measured insome laboratory experiments where spiked sediment has been aged for several days tomonths (Fu, Kan, & Tomson, 1994; Hatzinger & Alexander, 1995; Kan, Fu, & Tomson,1994; Ogram et al., 1985). However, we found that when a phenanthrene spike is incu-bated with sediment for 1–2 d, desorption of phenanthrene was reversible and well pre-dicted by the model over a 6 day period (LeBlanc, 2001).

The first-order desorption rate constant (an approximation of the spherical diffusionmodel), k (s�1) is given by Chen (1993)

k ¼ b� Deff=R2;

where b = 10.56Kd · MS/Vw + 22.7, and Deff ¼ Dm � n2=ð1� nÞ � rs � KD

Deff (cm2 s�1): an effective diffusion coefficient retarded by sorption,Dm (cm2 s�1): the molecular diffusion coefficient for dissolved phenanthrene,n: the intra-aggregate porosity,


rs (g cm3): the density of the solid phase, and

R2 (cm): the average radius of the sediment aggregate.

Assuming a value of 4.36 · 10�6 cm2/s for Dm (calculated from Hayduk & Laudie,1974, in Lyman, Reehl, & Rosenblatt, 1990), a typical value of 0.13 for n (Wu & Gsch-wend, 1986), a predicted value of 690 for the KD of phenanthrene (Schwarzenbachet al., 1993) and 10�2 cm for R, a desorption rate constant (k) of 2.3 · 10�3 s�1 can be esti-mated. The value of 10�2 cm for aggregate radius (R) is likely an overestimate. A 10�3 cmradius would result in a 100-fold increase in the predicted desorption rate constant. Com-paring k (=2.3 · 10�3 s�1) to the measured mineralization rate constants for bulk sedi-ments (4.1 · 10�7 s�1 and 1.0 · 10�7 s�1 for 9.6 and 12 d�1 treatments, respectively,converted from d�1 to s�1), suggests that mineralization rates for phenanthrene are severalorders of magnitude slower than desorption rates. However, if microbial uptake is onlyfrom pore waters, the microbial degradation rate should be normalized to the fractionof the phenanthrene substrate that is in pore waters. Utilizing an assumption of local equi-librium to obtain a dissolved pore water concentration, the resulting observed degradationrates for pore water phenanthrene were at most 50 times slower than the estimated desorp-tion rate (4.2 · 10�5 s�1 for 12 d�1). Thus even in pore waters, predicted desorption ratesappeared to be at least an order of magnitude faster than biological reaction rates. This isconsistent with the assumption of local equilibria and supports our finding that desorptionrate is not limiting to mineralization rate for easily desorbable phenanthrene spiked intosediments and equilibrated for short periods of time. Compound bioavailability, whichhas been shown to limit PAH degradation in natural sediment–porewater systems, istherefore discounted as the primary reason for the differences observed in degradationrates between resuspension treatments. However, given that sediments with greater resus-pension frequencies had greater loss to the aqueous phase, bioavailability effects cannot becompletely discounted.

4.3. Other effects of resuspension on microbial degradation rates

Incubation of sediment in the flow-through microcosms appears to have stimulated aPAH-degrading bacterial community, although from this experiment it cannot be deter-mined whether the stimulation of the PAH-degrading bacterial populations results in anincrease in physiological response, bacterial numbers, or both. The experiment in whichnative sediments were subjected to 0 and 12 d�1 resuspension frequencies for 30 days priorto incubation with 14C-labeled phenanthrene showed pronounced differences in 14C-phen-anthrene degradation rates under the same oxygen exposure conditions (Table 1). Mea-surements of ATP and bacterial cell counts in resuspended sediments from other flow-through experiments using our system showed greater biomass in resuspended sedimentswhen compared to sediments which were not resuspended (Gulnick, 2000). Findlay, Trex-ler, Guckert, et al. (1990) and Findlay, Trexler and White (1990) found increased hetero-trophic biomass and activity in sediments disturbed by physical and biological processes.Simultaneous increases in heterotrophic bacterial activity and phenanthrene-degradingactivity in the presence of capitellid worms have been reported (Bauer, Kerr, Bautista,Decker, & Capone, 1988). However, the mechanisms by which PAH-degrading communi-ties are affected by biologically and physically-mediated disturbances are not clear. Onepossible mechanism for enhanced phenanthrene degradation in frequently resuspended


treatments would be greater relative loss of labile sediment organic matter, leading tochange in substrate preference towards more refractory PAHs.

4.4. Fate of phenanthrene metabolites

Characterization of aqueous extracts by TLC showed that phenanthrene metaboliteswere present in the water column throughout the experiment and accounted for up to25% of the activity in the aqueous phase over the first 11 days. Radiolabel in the watercolumn over this time accounted for only 4% of the total activity. It was not determinedin this study whether these transformed products originated in the water column (frombacteria on suspended particulates) or in the sediment bed (porewaters) and transferredto the water column through diffusive and turbulent exchange. It is interesting to note,however, that transformation products made up a higher percentage of aqueous activityin treatments with resuspended sediments (22%) than in undisturbed sediments (5%) onday 3. Near the end of the experiment (days 26 and 30), most of the dissolved label wasparent compound in all treatments (3–7% metabolites). McElroy, Tripp, Farrington,and Teal (1987) and McElroy et al. (1994) found that a significant percent of the activityin the water column (greater than 50%) was transformed PAH, as opposed to the originalspiked 14C-benz[a]anthracene. They concluded that the bulk of the metabolism occurredeither on suspended particulates or at the sediment–water interface.

Metabolites were found in sediments in this study (up to 30% of the original label), andwere detectable up to 30 days. The metabolite fractions were higher than reported byHeitkamp and Cerniglia (1987) who found 1–3% of the extractable naphthalene as metab-olites after one week, but similar to Hinga and Pilson (1987) who recovered up to 14%extractable 14C-labeled benz[a]anthracene in mesocosm sediments as metabolites after200 days. Other studies (McElroy, Farrington, & Teal, 1990; McElroy et al., 1994; Stein,Hom, Casillas, Friedman, & Varanasi, 1987) that had actively metabolizing worms or fishin the sediments, reported primarily parent compounds over relatively short term (7 and25 days) exposures. However, McElroy et al. (1987) found significant quantities of polarmetabolites in sediment microcosms, which persisted over 42 days. McElroy et al. (1990)hypothesized that greater microbial metabolism in the earlier experiments lead to transfor-mation products that persisted in the sediments, as opposed to the dominance of metazoanmetabolism seen in the later studies which resulted in more water-soluble products. Thefate of bacterial- and metazoan-produced metabolites in sedimentary systems remainsan area for future research.

The nature of the non-extractable or residual pool is unknown, but 14C-incorporatedinto a residual pool was produced by one of three scenarios. The first is that untrans-formed phenanthrene accessed binding sites within the sediment aggregates and becameresistant to solvent extraction. Aging of spiked PAH compounds (including phenanthrene)has been shown to lead to increased resistance to desorption and mild solvent extractionover monthly timescales (Fu et al., 1994; Hatzinger & Alexander, 1995; Kan et al., 1994;White, Kelsey, Hatzinger, & Alexander, 1997). It should be noted that the solvent extrac-tion technique used in this study was relatively robust, involving multiple solvents andextraction by sonication, making it unlikely that up to 30% of parent compound wouldbe unrecoverable. The second possibility is that the formation of the resistant phase wasrelated to bacterial transformation of phenanthrene. This may cause transformation prod-ucts to become complexed with the sediment organic material through covalent and/or


non-covalent bonding and become resistant to base neutral solvent extraction. A thirdpossibility is that transformed label was incorporated into bacterial biomass and didnot partition into the solvent during extraction. The latter two scenarios seem to be morelikely, based upon two pieces of evidence. The first is that there was more label incorpo-ration into the residual pool with greater phenanthrene-degrading activity, as measured bymineralization (Fig. 5(b)). The second line of evidence was that in other resuspensionexperiments (discussed in Gulnick, 2000), killed controls yielded 0% recovery of label inthe residual phase. Also, in experiments with benzo[a]pyrene, where mineralization wasvery low (0.8–11% of initial label added after 89 days), there was a lower percentage ofthe initial label associated with the residual pool (2–12%, Gulnick, 2000).

4.5. Significance to field situations

The purpose of this study was to examine the effect of increased oxygenation throughsediment resuspension on PAH degradation, as well as to uncover other effects due toresuspending sediment. As such, the microcosms did not represent field situations wheresorption onto highly retentive phases can limit degradation, potentially overwhelmingthe resuspension effects demonstrated here. However, readily available hydrocarbons doexist in nature, as the result of fresh inputs due to oil spills, storm runoff, wastewater treat-ment plant and combined sewer overflow discharges (Hoffman, Mills, Latimer, & Quinn,1984; Neff, 1979; Van Vleet & Quinn, 1978).

This study provides compelling evidence for the influence of sediment resuspension onenhancing bacterial activity, and may aid in the interpretation of chemical concentrationdata obtained from areas where resuspension occurs on a large scale (such as storm ortidal-induced resuspension) or on a micro scale (such as the influence of bioturbating ben-thic macrofauna). This study also demonstrates the importance of oxygen for degradingbioavailable PAH compounds. Remediation efforts for PAH-contaminated sedimentsshould include enhanced oxygenation in addition to bacterial inocula and alternative elec-tron acceptors.

5. Summary

Suboxic sediments labeled with 14C-phenanthrene and resuspended at various frequen-cies into overlying waters in flow-through microcosms exhibited increased rates andextents of phenanthrene mineralization with increased resuspension frequencies. To a firstapproximation mineralization rates could be modeled as a first order process dependentupon the degree of oxygenation, which varied as a function of resuspension frequency.Metabolites of phenanthrene were detected in sediments even after 30 days with frequentresuspension. Although incorporation of phenanthrene into sediment bacteria was notmeasured directly, the accumulation of sediment-bound radiolabel in a solvent non-extractable pool may represent label that had been incorporated into bacterial biomass.Another possibility is that bacterially-transformed radiolabel complexed with sedimentmacromolecular organic matter. These studies strongly suggest that the mineralizationrate of phenanthrene in spiked sediments increases as a function of degree of oxygenationand is not limited by desorption. The fact that microbial degradation in these bedded flow-through experiments could be modeled with mineralization rates determined in separateslurry bottle studies with different sediment: water ratios is of interest.


Acknowledgments

The authors acknowledge the help of Judy Nichols in the laboratory and Tom Wilsonof the MSRC electronics shop for overall construction of the flow-through microcosm sys-tem. This research was funded by the US Office of Naval Research, grant no. R&T4110005 meq and a doctoral fellowship from the Hudson River Foundation.

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The influence of sediment resuspension on the degradation of phenanthrene in flow-through microcosms

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