Kerstin E. Scherr

Changes in iso- and n-alkane distribution during biodegradation of crude oil under nitrate and sulphate reducing conditions

Crude oil consists of a large number of hydrocarbons with different susceptibility to microbial degradation. The influence of hydrocarbon structure and molecular weight on hydrocarbon biodegradation under anaerobic conditions is not fully explored. In this study oxygen, nitrate and sulphate served as terminal electron acceptors (TEAs) for the microbial degradation of a paraffin-rich crude oil in a freshly contaminated soil. During 185 days of incubation, alkanes from n-C11 to n-C39, three n- to iso-alkane ratios commonly used as weathering indicators and the unresolved complex mixture (UCM) were quantified and statistically analyzed. The use of different TEAs for hydrocarbon degradation resulted in dissimilar degradative patterns for n- and iso-alkanes. While n-alkane biodegradation followed well-established patterns under aerobic conditions, lower molecular weight alkanes were found to be more recalcitrant than mid- to high-molecular weight alkanes under nitrate-reducing conditions. Biodegradation with sulphate as the TEA was most pronounced for long-chain (n-C32 to n-C39) alkanes. The observation of increasing ratios of n-C17 to pristane and of n-C18 to phytane provides first evidence of the preferential degradation of branched over normal alkanes under sulphate reducing conditions. The formation of distinctly different n- and iso-alkane biodegradation fingerprints under different electron accepting conditions may be used to assess the occurrence of specific degradation processes at a contaminated site. The use of n- to iso-alkane ratios for this purpose may require adjustment if applied for anaerobic sites.

Changes in bacterial communities from anaerobic digesters during petroleum hydrocarbon degradation

Anaerobic biodegradation of petroleum hydrocarbons (PHC) to methane has been recognized to occur in oil reservoirs and contaminated surface sites alike. This process could be employed efficiently for the treatment of contaminated materials, including petrochemical wastes and PHC-contaminated soil, since no external electron acceptor is required. Moreover, the controlled production of methane in digestion plants, similarly to the anaerobic digestion (AD) of energy crops or organic residues, would enable for energy recovery from these wastes. At present, little is known about the bacterial communities involved in and responsible for hydrocarbon fermentation, the initial step in PHC conversion to methane. In the present study, the fate of two different methanogenic communities derived from the AD of wastewater (WWT) and of biowaste, mixed with PHC-contaminated soil (SWT), was monitored during incubation with PHC using denaturing gradient gel electrophoresis (DGGE) of 16S rDNA genes amplified with Bacteria-specific primers. During 11 months of incubation, slight but significant degradation of PHC occurred in both sludges and distinct bacterial communities were developing. In both sludges, Bacteroidetes were found. In addition, in WWT, the bacterial community was found to be dominated by Synergistetes and Proteobacteria, while Firmicutes and unidentified members were abundant in SWT. These results indicate that bacterial communities from anaerobic digesters can adapt to and degrade petroleum hydrocarbons. The decontamination of PHC-containing waste via fermentative treatment appears possible.

Influence of the nature of soil organic matter on the sorption behaviour of pentadecane as determined by PLS analysis of mid-infrared DRIFT and solid-state 13C NMR spectra

The nature of soil organic matter (SOM) functional groups associated with sorption processes was determined by correlating partitioning coefficients with solid-state (13)C nuclear magnetic resonance (NMR) and diffuse reflectance mid-infrared (DRIFT) spectral features using partial least squares (PLS) regression analysis. Partitioning sorption coefficients for n-pentadecane (n-C(15)) were determined for three alternative models: the Langmuir model, the dual distributed reactive domain model (DRDM) and the Freundlich model, where the latter was found to be the most appropriate. NMR-derived constitutional descriptors did not correlate with Freundlich model parameters. By contrast, PLS analysis revealed the most likely nature of the functional groups in SOM associated with n-C(15) sorption coefficients (K(F)) to be aromatic, possibly porous soil char, rather than aliphatic organic components for the presently investigated soils. High PLS cross-validation correlation suggested that the model was robust for the purpose of characterising the functional group chemistry important for n-C(15) sorption.

Sequential application of electron donors and humic acids for the anaerobic bioremediation of chlorinated aliphatic hydrocarbons.

In situ anaerobic bioremediation of chlorinated solvents such as perchloroethene (PCE) frequently faces the problem of accumulating toxic, lower chlorinated compounds such as dichloroethene (cis-DCE) and vinyl chloride (VC). In the present study, the efficacy of the sequential application of electron donors, supporting reductive dechlorination, and of humic acids, acting as extracellular electron shuttles facilitating the anaerobic oxidation of recalcitrant intermediates, was explored in microcosm studies. Upon one initial dose of lactose, supplied in a 1000-fold superstoichiometric electron equivalent ratio, PCE was completely converted into cis-DCE within 35 days. Repeated electron donor additions did not entail exhaustive cis-DCE degradation over incubation time (120 days). Although the electron donor was quickly converted into fatty acids, about 30% of added reducing equivalents were recovered as acetate after four months of operation, indicating the inhibition of acetoclastic methanogenesis. In the next step, the substoichiometric addition of anthraquinone-2,6-disulfonate, a humic acid model compound, effected the complete removal of the accumulated cis-DCE within 15 days, probably as a result of the participation of the quinone in the biotic or abiotic anaerobic oxidation of cis-DCE. Cis-DCE degradation was not connected to the accumulation of VC, rendering the proposed two-step treatment an efficient and environmentally compliant remedy for anaerobic groundwater bodies contaminated with chlorinated solvents.

Comprehensive GC2/MS for the monitoring of aromatic tar oil constituents during biodegradation in a historically contaminated soil

The constituents of tar oil comprise a wide range of physico-chemically heterogeneous pollutants of environmental concern. Besides the sixteen polycyclic aromatic hydrocarbons defined as priority pollutants by the US-EPA (EPA-PAHs), a wide range of substituted (NSO-PAC) and alkylated (alkyl-PAC) aromatic tar oil compounds are gaining increased attention for their toxic, carcinogenic, mutagenic and/or teratogenic properties. Investigations on tar oil biodegradation in soil are in part hampered by the absence of an efficient analytical tool for the simultaneous analysis of this wide range of compounds with dissimilar analytical properties. Therefore, the present study sets out to explore the applicability of comprehensive two-dimensional gas chromatography (GC²/MS) for the simultaneous measurement of compounds with differing polarity or that are co-eluting in one-dimensional systems. Aerobic tar oil biodegradation in a historically contaminated soil was analyzed over 56 days in lab-scale bioslurry tests. Forty-three aromatic compounds were identified with GC²/MS in one single analysis. The number of alkyl chains on a molecule was found to prime over alkyl chain length in hampering compound biodegradation. In most cases, substitution of carbon with nitrogen and oxygen was related to increased compound degradation in comparison to unalkylated and sulphur- or unsubstituted PAH with a similar ring number.The obtained results indicate that GC²/MS can be employed for the rapid assessment of a large variety of structurally heterogeneous environmental contaminants. Its application can contribute to facilitate site assessment, development and control of microbial cleanup technologies for tar oil contaminated sites.

Distillation fraction-specific ecotoxicological evaluation of a paraffin-rich crude oil

Crude oil is a complex mixture of petroleum hydrocarbons (PHC) with distinct chemical, physical and toxicological properties relevant for contaminated site risk assessment. Ecotoxicological effects of crude oil distillation fractions on luminescent bacteria (Vibrio fischeri), earthworms (Dendrobaena hortensis) and invertebrates (Heterocypris incongruens) were tested using two spiked soils and their elutriates. Fraction 2 (F2) had an equivalent carbon number (ECN) range of >10 to 16, and F3 from >16 to 39. F2 showed a substantially higher ecotoxicological effect than F3 for Vibrio and Dendrobaena. In contrast, severe inhibition of Heterocypris by the poorly soluble F3 is attributed to mechanical organ blockage. Immediate sequestration of PHC to the organic matter-rich soil effected reduced toxicity for all organisms. This study indicates that a more differentiated consideration (i) of PHC mixtures based on ECN range and (ii) of model soil properties employed for ecotoxicity testing should be included into PHC-contaminated site risk assessment.

Occurrence of Chlorotriazine herbicides and their transformation products in arable soils

Chlorotriazine herbicides (CTs) are widely used pest control chemicals. In contrast to groundwater contamination, little attention has been given to the circumstances of residue formation of parent compounds and transformation products in soils. Seventy-five cultivated floodplain topsoils in the Czech Republic were sampled in early spring of 2015, corresponding to a minimum of six months (current-use terbuthylazine, TBA) and a up to a decade (banned atrazine, AT and simazine, SIM) after the last herbicide application. Soil residues of parent compounds and nine transformation products were quantified via multiple residue analysis using liquid chromatography - tandem mass spectrometry of acetonitrile partitioning extracts (QuEChERS). Using principal component analysis (PCA), their relation to soil chemistry, crops and environmental parameters was determined. Of the parent compounds, only TBA was present in more than one sample. In contrast, at least one CT transformation product, particularly hydroxylated CTs, was detected in 89% of the sites, or 54% for banned triazines. Deethylated and bi-dealkylated SIM or AT residues were not detectable. PCA suggests the formation and/or retention of CT hydroxy-metabolite residues to be related to low soil pH, and a direct relation between TBA and soil organic carbon, and between deethyl-TBA and clay or Ca contents, respectively, the latter pointing towards distinct sorption mechanisms. The low historic application of simazine contrasted by the high abundance of its residues, and the co-occurrence with AT residues suggests the post-ban application of AT and SIM banned triazines as a permitted impurity of TBA formulations as a recent, secondary source. The present data indicate that topsoils do not contain abundant extractable residues of banned parent chlorotriazines, and are thus likely not the current source for related ground- and surface water contamination. In contrast, topsoils might pose a long-term source of TBA and CT transformation products for ground and surface water contamination.

Extracellular Electron Transfer in in situ Petroleum Hydrocarbon Bioremediation

Anthropogenic contamination of soil, subsurface and surface waters and atmosphere with toxic organic chemicals is an environmental issue of world wide concern. While energy and goods production from fossil hydrocarbon sources is one of the driving factors of global economy, the often adverse environmental effects of exploration, production, transport and processing of crude and shale oils, tar sands, coal and natural gas seldom come into the focus of attention.

Composition and Dissolution of a Migratory, Weathered Coal Tar Creosote DNAPL

Opaque, viscous tars derived from the carbonization of fossile carbon feedstocks, such coal tars and creosote, are long-term sources of groundwater contamination, predominantly with poly- and heterocyclic aromatic hydrocarbons (PAH). The dissolution, ageing and migratory behavior of dense, non aqueous phase liquid (DNAPL) coal tar blobs and pools forming at the aquitard is not sufficiently understood to estimate the risk and adequately design groundwater treatment measures at a contaminated site. In this study, we investigate the composition and dissolution of a migrated, aged creosote DNAPL and corresponding experimental and groundwater profiles using comprehensive two-dimensional gas chromatography (GCxGC-MS). GC-FID unresolved compounds were attributed to methylated homocyclic species using GCxGC-MS in the Methylanthracene weight range. Equilibrium concentrations were estimated using Raoult’s law, assuming non-ideal behavior. Low molecular weight compounds were found to be prevalent even after decades of weathering, with Naphthalene (8% by mass) representing the most abundant identified compound, contrary to the expected preferential depletion of hydrophilic compounds. Morevoer, dimethylnaphthalenes were relatively more abundant in the aqueous boundary layer than in the DNAPL. DNAPL migration over 400m with the groundwater flow effected lower viscosity and specific gravity of the migrated phase body in a superposition of weathering, transport and aquifer chromatography effects. Based on a decomposition of analysed and estimated constituents using the group contribution approach, reference DNAPL values for activity coefficients γi were used to model aqueous solubilities for selected compounds. Anthracene was close to its theoretical precipitation limit in the bulk DNAPL. While laboratory and modelled DNAPL dissolution behavior agree well, field data imply the presence of specific interfacial in situ processes significantly impacting dissolution processes. Based on aqueous GCxGC-MS profiles over the DNAPL, a hypothetical interfacial in situ film was calculated to be composed primarily of Phenanthrene, with minor contribution by Naphthalene, possibly forming a viscous barrier for the dissolution of lower molecular weight PAH. The main advances and gaps in electron donor DNAPL understanding are discussed regarding our conception of weathered, migrating hydrophobic DNAPL bodies in the aquifer of historic contaminated sites for the adequate treatment of contaminated water.