Methodology for the Determination of Rates of Microbial Transformation of Polycyclic Aromatic Hydrocarbons in Sediments

    Published: Jan 1979

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    A methodology has been developed for measurement of aerobic rates of microbial transformation of polycyclic aromatic hydrocarbons (PAH) in sediment samples. Individual 14C-labeled PAH are added to sediment aliquots in sealed vials containing potassium hydroxide (KOH) traps. Vials containing sterile sediment portions are treated similarly and serve as controls. Sediment microbial activity is halted at intervals by addition of acetone to each vial. Following quantitation of 14CO2 in each trap by liquid scintillation counting, each sediment portion is extracted with acetone in a micro-Soxhlet apparatus. Extractable 14C is separated into unaltered 14C-PAH and polar transformation products by column or thin-layer chromatography; each fraction is quantified by liquid scintillation or automated thin-layer scanning. Residual 14C is measured by combustion of extracted sediment and liquid scintillation counting of the 14CO2 evolved.

    Tests show that the presence of acetone in amounts used in the assay does not significantly alter transformation rates. The measured rate of anthracene transformation was proportional to 14C-anthracene concentration at the substrate level employed, thus suggesting that addition of 14C-labeled substrate does not alter significantly the microbial transformation rate. To date sediments from three locations have been examined: a pristine stream, one receiving runoff from a petroleum storage depot, and a stream receiving a byproduct coking plant effluent discharge. Transformations of 14C-naphthalene, anthracene, and benz(a)anthracene have been observed in the two contaminated streams. Evolution of 14CO2 as a proportion of substrate added varies greatly between compounds: while 64 percent of 14C-naphthalene transformed during one assay was trapped in KOH, corresponding values for 14C-anthracene and 14C-benz(a)anthracene were 11 and 15 percent, respectively. The principal transformation product in most assays is residual 14C, which presumably has been incorporated into cells in a nonextractable form. Polar extractable metabolites are also quantitatively significant, particularly for 14C-anthracene and 14C-benz(a)anthracene, although thin-layer chromatographic patterns of metabolite formation from 14C-anthracene differ between sediments obtained from waters receiving coking plant effluent and petroleum storage runoff. The assay procedure permits a more valid estimation of microbial transformation rates in sediment than do procedures previously applied.


    microbial, degradation, sediment, chromatography, transformation, rate, polycyclic aromatic hydrocarbon

    Author Information:

    Schwall, LR
    Consultant and research associate, Oak,

    Herbes, SE
    Consultant and research associate, Oak,

    Paper ID: STP38149S

    Committee/Subcommittee: D19.24

    DOI: 10.1520/STP38149S

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