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A reliance on diesel generated power and a history of imperfect fuel management have created a legacy of petroleum hydrocarbon contamination at subantarctic Macquarie Island. Increasing environmental awareness and advances in contaminant characterisation and remediation technology have fostered an impetus to reduce the environmental risk associated with legacy sites. A funnel and gate permeable bio-reactive barrier (PRB) was installed in 2014 to address the migration of Special Antarctic Blend diesel from a spill that occurred in 2002, as well as older spills and residual contaminants in the soil at the Main Power House. The PRB gate comprised of granular activated carbon and natural clinoptilolite zeolite. Petroleum hydrocarbons migrating in the soil water were successfully captured on the reactive materials, with concentrations at the outflow of the barrier recorded as being below reporting limits. The nutrient and iron concentrations delivered to the barrier demonstrated high temporal variability with significant iron precipitation observed across the bed. The surface of the granular activated carbon was largely free from cell attachment while natural zeolite demonstrated patchy biofilm formation after 15 months following PRB installation. This study illustrates the importance of informed material selection at field scale to ensure that adsorption and biodegradation processes are utilised to manage the environmental risk associated with petroleum hydrocarbon spills. This study reports the first installation of a permeable bio-reactive barrier in the subantarctic., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Nitrogen deficiency has been identified as the main inhibiting factor for biodegradation of petroleum hydrocarbons in low nutrient environments. This study examines the performance of ammonium exchanged zeolite to enhance biodegradation of petroleum hydrocarbons migrating in soil water within laboratory scale flow cells. Biofilm formation and biodegradation were accelerated by the exchange of cations in soil water with ammonium in the pores of the exchanged zeolite when compared with natural zeolite flow cells. These results have implications for sequenced permeable reactive barrier design and the longevity of media performance within such barriers at petroleum hydrocarbon contaminated sites deficient in essential soil nutrients., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The application of controlled release nutrient (CRN) materials to permeable reactive barriers to promote biodegradation of petroleum hydrocarbons in groundwater was investigated. The longevity of release, influence of flow velocity and petroleum hydrocarbon concentration on nutrient release was assessed using soluble and ion exchange CRN materials; namely Polyon™ and Zeopro™. Both CRN materials, assessed at 4 °C and 23 °C, demonstrated continuing release of nitrogen, phosphorus and potassium (N-P-K) at 3500 bed volumes passing, with longer timeframes of N-P-K release at 4 °C. Zeopro™-activated carbon mixtures demonstrated depletion of N-P-K prior to 3500 bed volumes passing. Increased flow velocity was shown to lower nutrient concentrations in Polyon™ flow cells while nutrient release from Zeopro™ was largely unchanged. The presence of petroleum hydrocarbons, at 1.08 mmol/L and 3.25 mmol/L toluene, were not shown to alter nutrient release from Polyon™ and Zeopro™ across 14 days. These findings suggest that Polyon™ and Zeopro™ may be suitable CRN materials for application to PRBs in low nutrient environments., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

A field trial was conducted at Casey Station, Antarctica to assess the suitability of a permeable reactive barrier (PRB) media sequence for the remediation of sites containing both hydrocarbon and heavy metal contamination. An existing PRB was modified to assess a sequence consisting of three sections: (i) Nutrient release/hydrocarbon sorption using ZeoPro™ and granular activated carbon; (ii) Phosphorus and heavy metal capture by granular iron and sand; (iii) Nutrient and excess iron capture by zeolite. The media sequence achieved a greater phosphorus removal capacity than previous Antarctic PRB configurations installed on site. Phosphorus concentrations were reduced during flow through the iron/sand section and iron concentrations were reduced within the zeolite section. However, non-ideal flow was detected during a tracer test and supported by analysis of media and liquid samples from the second summer of operation. Results indicate that the PRB media sequence trialled might be appropriate for other locations, especially less environmentally challenging contaminated sites., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Bioremediation has been used to remediate petroleum hydrocarbon (PHC)-contaminated sites in polar regions; however, limited knowledge exists in understanding how frozen conditions influence factors that regulate microbial activity. We hypothesized that increased liquid water (θ(liquid) ) would affect nutrient supply rates (NSR) and gas diffusion under frozen conditions. If true, management practices that increase θ(liquid) should also increase bioremediation in polar soils by reducing nutrient and oxygen limitations. Influence of θ(liquid) on NSR was determined using diesel-contaminated soil (0-8,000 mg kg(-1)) from Casey Station, Antarctica. The θ(liquid) was altered between 0.007 and 0.035 cm(3) cm(-3) by packing soil cores at different bulk densities. The nutrient supply rate of NH 4+ and NO 3-, as well as gas diffusion coefficient, D(s), were measured at two temperatures, 21°C and -5°C, to correct for bulk density effects. Freezing decreased NSR of both NH 4+ and NO 3-, with θ(liquid) linked to nitrate and ammonia NSR in frozen soil. Similarly for D(s), decreases due to freezing were much more pronounced in soils with low θ(liquid) compared to soils with higher θ(liquid) contents. Additional studies are needed to determine the relationship between degradation rates and θ(liquid) under frozen conditions., (Copyright © 2011 SETAC.)

Potential microbial activities are commonly used to assess soil toxicity of petroleum hydrocarbons (PHC) and are assumed to be a surrogate for microbial activity within the soil ecosystem. However, this assumption needs to be evaluated for frozen soil, in which microbial activity is limited by liquid water (θ(liquid)). Influence of θ(liquid) on in situ toxicity was evaluated and compared to the toxicity endpoints of potential microbial activities using soil from an aged diesel fuel spill at Casey Station, East Antarctica. To determine in situ toxicity, gross mineralization and nitrification rates were determined by the stable isotope dilution technique. Petroleum hydrocarbon-contaminated soil (0-8,000 mg kg(-1)), packed at bulk densities of 1.4, 1.7, and 2.0 g cm(-3) to manipulate liquid water content, was incubated at -5°C for one, two, and three months. Although θ(liquid) did not have a significant effect on gross mineralization or nitrification, gross nitrification was sensitive to PHC contamination, with toxicity decreasing over time. In contrast, gross mineralization was not sensitive to PHC contamination. Toxic response of gross nitrification was comparable to potential nitrification activity (PNA) with similar EC25 (effective concentration causing a 25% effect in the test population) values determined by both measurement endpoints (400 mg kg(-1) for gross nitrification compared to 200 mg kg(-1) for PNA), indicating that potential microbial activity assays are good surrogates for in situ toxicity of PHC contamination in polar regions., (Copyright © 2011 SETAC.)

We do not yet understand why fuel spills can cause greater damage in polar soils than in temperate soils. The role of water in the freezing environment may partly be responsible for why polar soils are more sensitive to pollution. We hypothesized that hydrocarbons alter the liquid water in frozen soil, and we evaluated this hypothesis by conducting laboratory and field experiments at Casey Station, Antarctica. Liquid water content in frozen soils (theta(liquid)) was estimated by time domain reflectometry in laboratory, field collected soils, and in situ field measurements. Our results demonstrate an increase in liquid water associated with hydrocarbon contamination in frozen soils. The dependence of theta(liquid) on aged fuel and spiked fuel were almost identical,with a slope of 2.6 x 10(-6) mg TPH (total petroleum hydrocarbons) kg(-1) for aged fuel and 3.1 x 10(-6) mg TPH kg(-1) for spiked fuel. In situ measurements found theta(liquid) depends, r2 = 0.75, on fuel for silt loam soils (theta(liquid) = 0.094 + 7.8 x 10(-6) mg TPH kg(-1)) but not on fuel for silt clay loam soils. In our study, theta(liquid) doubled in field soils and quadrupled in laboratory soils contaminated with diesel which may have profound implications on frost heave models in contaminated soils.

Sub-Antarctic islands have been subjected to petroleum hydrocarbon spills, yet no information is available regarding the toxicity of petroleum hydrocarbons to these subpolar soils. The purpose of the present study was to identify soil biogeochemical toxicity end points for petroleum hydrocarbon contamination in sub-Antarctic soil. Soil from Macquarie Island, a sub-Antarctic island south of Australia, was collected and exposed to 10 concentrations of Special Antarctic Blend (SAB) diesel fuel, ranging from 0 to 50,000 mg fuel/kg soil, for a 21-d period. The sensitivity of nitrification, denitrification, carbohydrate utilization, and total soil respiration to SAB fuel was assessed. Potential nitrification activity was the most sensitive indicator of SAB contamination assessed for nitrogen cycling, with an IC20 (concentration that results in a 20% change from the control response) of 190 mg fuel/ kg soil. Potential denitrification activity was not as sensitive to SAB contamination, with an IC20 of 950 mg fuel/kg soil for nitrous oxide production. Nitrous oxide consumption was unaffected by SAB contamination. Carbohydrate utilization (respiration caused by sucrose) was a more sensitive indicator (IC20, 16 mg fuel/kg soil) of SAB contamination than total respiration (IC20, 220 mg fuel/kg soil). However, total soil respiration was a more responsive measurement end point, increasing soil respiration over a 72-h period by 17 mg of CO2, compared to a change of only 2.1 mg of CO2 for carbohydrate utilization. Our results indicate that IC20s varied between 16 to 950 mg fuel/kg soil for Macquarie Island soil spiked with SAB diesel fuel. These results indicate that current cleanup levels derived from temperate zones may be too liberal for soil contamination in sub-Antarctic islands.

A real-time polymerase chain reaction (PCR) method to quantify the proportion of microorganisms containing alkane monooxygenase was developed and used to follow changes in the microbial community in hydrocarbon-contaminated Antarctic soil during a bioremediation field trial. Assays for the alkB and rpoB genes were validated and found to be both sensitive and reproducible (less than 2% intrarun variation and 25-38% interrun variation). Results from the real-time PCR analysis were compared to analysis of the microbial population by a culture-based technique [most probable number (MPN) counts]. Both types of analysis indicated that fertilizer addition to hydrocarbon-contaminated soil stimulated the indigenous bacterial population within 1 year. The proportion of alkB containing microorganisms was positively correlated to the concentration of n-alkanes in the soil. After the concentration of n-alkanes in the soil decreased, the proportion of alkane-degrading microorganisms decreased, but the proportion of total hydrocarbon-degrading microorganisms increased, indicating another shift in the microbial community structure and ongoing biodegradation.

Human activities in the Antarctic have resulted in hydrocarbon contamination of these fragile polar soils. Bioremediation is one of the options for remediation of these sites. However, little is known about anaerobic hydrocarbon degradation in polar soils and the influence of bioremediation practices on these processes. Using a field trial at Old Casey Station, Antarctica, we assessed the influence of fertilization on the anaerobic degradation of a 20-year old fuel spill. Fertilization increased hydrocarbon degradation in both anaerobic and aerobic soils when compared to controls, but was of most benefit for anaerobic soils where evaporation was negligible. This increased biodegradation in the anaerobic soils corresponded with a shift in the denitrifier community composition and an increased abundance of denitrifiers and benzoyl-CoA reductase. A microcosm study using toluene and hexadecane confirmed the degradative capacity within these soils under anaerobic conditions. It was observed that fertilized anaerobic soil degraded more of this hydrocarbon spike when incubated anaerobically than when incubated aerobically. We conclude that denitrifiers are actively involved in hydrocarbon degradation in Antarctic soils and that fertilization is an effective means of stimulating their activity. Further, when communities stimulated to degrade hydrocarbons under anaerobic conditions are exposed to oxygen, hydrocarbon degradation is suppressed. The commonly accepted belief that remediation of polar soils requires aeration needs to be reevaluated in light of this new data.

In many temperate regions, fuel and oil spills are sometimes managed simply by allowing natural degradation to occur, while monitoring soils and groundwater to ensure that there is no off-site migration or on-site impact. To critically assess whether this approach is suitable for coastal Antarctic sites, we investigated the extent of evaporation and biodegradation at three old fuel spills at Casey Station. Where the contaminants migrated across frozen ground, probably beneath snow, approximately half the fuel evaporated in the first few months prior to infiltration at the beginning of summer. Once in the ground, however, evaporation rates were negligible. In contrast, minor spills from fuel drums buried in an abandoned waste disposal site did not evaporate to the same extent. Biodegradation within all three spill sites is generally very minor. We conclude that natural attenuation is not a suitable management strategy for fuel-contaminated soils in Antarctic coastal regions.

A large-scale, in situ experiment was set up near the Bailey Peninsula area (Casey Station, East Antarctica) to monitor the natural attenuation of synthetic lubricants in marine sediments over five years. Here, we report the short-term changes after 5 and 56 weeks. The lubricants tested were an unused and used Mobil lubricant (0W/40; Exxon Mobil, Irving, TX, USA) and a biodegradable alternative (0W/20; Fuchs Lubricants, Harvey, IL, USA). Clean sediment was collected, contaminated with the lubricants, and deployed by divers onto the seabed in a randomized block design. The sampled sediments were analyzed by gas chromatography-flame-ionization detector and gas chromatography-mass spectrometry with selective ion monitoring. The base fluid of all lubricant treatments did not decrease significantly after 56 weeks in situ. Alkanoate esters of 1,1,1-tris(hydroxymethyl)propane in the biodegradable and unused lubricants were degraded extensively in situ; however, these esters constituted only a minor proportion of the lubricant volume. The additives, alkylated naphthalenes and substituted diphenylamines, were fairly resistant to degradation, which is of environmental concern because of their toxicity. The biodegradable lubricant did not break down to recognized biodegradable thresholds and, as such, should not be classified as biodegradable under Antarctic marine conditions. A separate experiment was conducted to determine the influence of sediment preparation and deployment on compound ratios within the lubricants, and we found that preparation and deployment of the contaminated sediments had only a minor effect on compound recovery. Further monitoring of this in situ experiment will provide much needed information about the long-term natural attenuation of lubricants.

A small-scale (<500 m length) transect-based survey was conducted in December 1998 to examine the spatial distribution of soft-sediment communities and of concentrations of heavy metals and hydrocarbons in sediments in Brown Bay, adjacent to an abandoned waste dump, at Casey Station, Antarctica. Samples were taken along three transects at increasing distances (nine stations) from the shore-line waste dump. A gradient of contamination was detected, but concentrations of contaminants were very variable with "hotspots" or high levels of contaminants at some stations. Multivariate analysis revealed that the distribution of soft-sediment communities was distinctly different between the inner, middle and outer stations. Abundances of most taxa were very variable with few patterns apparent, but some fauna displayed an abundance gradient from the inner to the outer part of the bay. Many taxa had maximum abundances at outer stations and minimum at inner stations. Multivariate correlations between environmental variables and soft-sediment communities indicated that combinations of some metals (Cd, Cu, Sn, Pb) and grain size (mainly finer fractions, fine sands and coarse silts) were the variables that best "matched" the community patterns within Brown Bay. This study indicated that there were significant correlations between the presence of contaminants and the distribution and composition of soft-sediment communities over very small spatial scales.

Abstract: Granular activated carbon (GAC) has been used as an adsorbent for hydrocarbons in a range of permeable reactive barriers. This work investigates the influence of temperature on adsorption performance. In particular, the influence of temperature in the range of 20°C to 4°C on the sorption equilibrium and kinetics of toluene on GAC surface were investigated. The results show that low temperature leads to decreased toluene sorption by GAC and slower reaction kinetics. Sorption kinetics studies show that diffusion coefficients are also lower at 4°C (3.65×10−13 m2 s−1) than 20°C (5.112×10−13 m2 s−1). [Copyright &y& Elsevier]

Fuel spills in Antarctica typically occur in rare ice-free oases along the coast, which are areas of extreme seasonal freezing. Spills often occur at subzero temperatures, but little is known of ecosystem sensitivity to pollutants, in particular the influence that soil liquid water and low temperature have on toxicity of petroleum hydrocarbons (PHC) in Antarctic soil. To evaluate PHC toxicity, 32 locations at an aged diesel spill site in Antarctica were sampled nine times to encompass frozen, thaw, and refreeze periods. Toxicity was assessed using potential activities of substrate-induced respiration, basal respiration, nitrification, denitrification, and metabolic quotient as well as microbial community composition and bacterial biomass. The most sensitive indicator was community composition with a PHC concentration effecting 25% of the population (EC25) of 800 mg/kg, followed by nitrification (2,000 mg/kg), microbial biomass (2,400 mg/kg), and soil respiration (3,500 mg/kg). Despite changes in potential microbial activities and composition over the frozen, thaw, and refreeze period, the sensitivity of these endpoints to PHC did not change with liquid water or temperature. However, the variability associated with ecotoxicity data increased at low liquid water contents. As a consequence of this variability, highly replicated (n =50) experiments are needed to quantify a 25% ecological impairment by PHCs in Antarctic soils at a 95% level of significance. Increases in biomass and respiration associated with changes in community composition suggest that PHC contamination in Antarctic soils may have irrevocable effects on the ecosystem. [ABSTRACT FROM AUTHOR]

Abstract: There is a common perception that terrestrial fuel spills in Antarctica are long lived and that plumes naturally degrade very slowly. However, previous studies have isolated hydrocarbon degrading microbes from many Antarctic soils and biodegradation has been documented at low temperatures in the laboratory. A detailed study of an old fuel spill at Casey Station, Antarctica, indicates that fuel has migrated down a small catchment into the marine environment. By integrating chemical signatures indicative of evaporation and biodegradation with environmental parameters it is possible to correlate natural attenuation and dispersal of the spill within the catchment. GIS terrain modelling techniques are used to explore the spatial patterns of biodegradation and evaporation in relation to slope, solar radiation, surface wetness, and landform properties. [Copyright &y& Elsevier]

Abstract: Little effort has been devoted to differentiating between hydrocarbon losses through evaporation and biodegradation in treatability studies of fuel-contaminated Antarctic soils. When natural attenuation is being considered as a treatment option, it is important to be able to identify the mechanism of hydrocarbon loss and demonstrate that rates of degradation are sufficient to prevent off-site migration. Similarly, where complex thermally enhanced bioremediation schemes involve nutrient addition, water management, air stripping and active heating, it is important to appreciate the relative roles of these mechanisms for cost minimisation. Following the loss of hydrocarbons by documenting changes in total petroleum hydrocarbons offers little insight into the relative contribution of evaporation and biodegradation. We present a methodology here that allows identification and quantification of evaporative losses of diesel range organics at a range of temperatures using successively less volatile compounds as fractionation markers. We also present data that supports the general utility of so-called biodegradation indices for tracking biodegradation progress. We are also able to show that at 4°C indigenous Antarctic soil bacteria degrade Special Antarctic Blend fuel components in the following order: naphthalene and methyl-napthalenes, light n-alkanes, then progressively heavier n-alkanes; whereas isoprenoids and the unresolved complex mixture are relatively recalcitrant. [Copyright &y& Elsevier]

A manipulative field experiment was conducted at Casey Station, Antarctica, to determine the effect of contamination of marine sediments on the recruitment and development of soft-sediment assemblages. Uncontaminated marine sediments were defaunated and then contaminated by adding hydrocarbons (diesel fuel and lubrication oil) or heavy metals (extracted from contaminated soil) prior to deployment on the seabed for a period of eleven weeks during an Antarctic summer (December to February) at three locations. Two of the locations were intended as reference locations (O''Brien and Sparkes Bay) and were well away from the influence of station activities, but Sparkes Bay was found to be naturally contaminated by some metals. The other location (Brown Bay) was adjacent to an abandoned waste disposal site and is contaminated by hydrocarbons and metals, and soft-sediment assemblages are different from those in control locations. Recruiting assemblages were significantly different between locations and there was a significant effect of sediment contamination. Assemblages in hydrocarbon contaminated sediment were significantly different to those in control and metal treated sediment at two of the locations (Brown and Sparkes Bay). There were no differences between assemblages in control sediment and metal-contaminated sediment. Univariate analyses (ANOVAs) indicated greater abundances of crustaceans, mainly due to several species of gammarid, in control and metal treatments than hydrocarbon-treated sediment. Further analysis (t-tests) revealed additional differences between hydrocarbon vs. both control and metal treatments, in the total abundance, and abundance of gammarids, tanaids, ostracods, isopods and copepods, particularly at Brown Bay and Sparkes Bay. There was also a trend of greater abundances of polychaetes and gastropods in hydrocarbon and metal treatments compared to the control. In comparison with the hydrocarbon treatment, the metal treatment had little effect. There was significant spatial variability in recruitment within each location, which may have obscured some effects of sediment contamination. There were also significant differences in variability of recruitment between locations. The rate of fauna recruiting to sediments in this experiment at Casey were greater than previously recorded in Antarctica. [Copyright &y& Elsevier]

Abstract: A series of nutrient-amended microcosms was used to investigate changing microbial communities during biodegradation in hydrocarbon-contaminated Antarctic soils at 4, 10 and 42 °C. Although sample heterogeneity resulted in no statistically significant reduction in total petroleum hydrocarbons, biodegradation ratios indicate significant mineralisation. The number of culturable bacterial grown at 4 and 10 °C increased from 3×105 g−1 dry soil then peaked after 5 days incubation at 5×107 g−1 dry soil, before decreasing to and remaining stable at ca. 2×107 g−1 dry soil. While the bacterial population grown at 42 °C was initially a minor constituent of the total culturable bacterial population, after 40 days there was similar numbers of bacteria estimated at all temperatures investigated. Denaturing gradient gel electrophoresis indicated significant differences in the microbial community between the 4 and 10 °C and the 42 °C microcosms. Numerically dominant culturable hydrocarbon degrading bacteria were isolated at each temperature; 16S rRNA gene sequences identified the 4 and 10 °C isolates as Pseudomonas spp., and the 42 °C isolates as Paenibacillus spp. Fatty acid methyl ester profiles of the cultures were consistent with these identifications. The results from this study indicate that bioremediation treatments will substantially alter the soil microbial ecosystem. All isolates from this study were capable of growth at 28 °C. The presence of autochthonous hydrocarbon degrading microbes capable of growth at higher temperatures introduces the possibility of in situ remediation treatment options involving heating (ca. 10–28 °C) to accelerate the rate of hydrocarbon degradation. [Copyright &y& Elsevier]

Abstract: In this investigation different sorbent materials (MYCELX coated sand, granular activated carbon (GAC) and surfactant-modifed zeolite (SMZ)) were tested for their suitability for capture of sparingly soluble hydrocarbons in soil water. These materials were assessed for their potential use in permeable reactive barriers in cold regions. Batch sorption tests at 20 °C and 4 °C were performed along with a number of surface characterization techniques, including thermogravimetric analysis, nitrogen adsorption and Fourier transform infrared analysis (FTIR). ionic strength and pH tests were also conducted to determine their impact on hydrocarbon sorption. The results of the surface characterization tests showed that the quality of the MYCELX coating on the sand was highly variable, thus leading to unreliable adsorption results. GAC proved to be the best sorbent material at high and low temperatures on a mass basis. However, on a surface area basis, SMZ performed better than GAC. High ionic strength and high and low pH had little effect for hydrocarbon sorption onto GAC and SMZ. Both GAC and SMZ exhibited reduced adsorption efficiency at 4 °C compared to 20 °C. The extent of the reduced adsorption efficiency varied depending on the hydrocarbon. [Copyright &y& Elsevier]

The Antarctic Treaty permits the discharge of wastewater into Antarctic marine waters providing that conditions exist for initial dilution and rapid dispersal. We investigated the dilution and dispersal of macerated wastewater around Australia's Davis Station in East Antarctica and examined sediments for evidence of contaminants. Methods used to examine hydrodynamic conditions included current meters, dye release experiments and measurement of sewage-associated microbial markers and surfactants in the water column. We measured marine sediments for metals, nutrients, PBDEs, hydrocarbons and faecal sterols. We propose that if there is adequate dilution and dispersal there would be no significant difference in contaminant concentrations in sediments around the outfall compared to distant control sites. Currents were strongly correlated with prevailing wind conditions. Modelling indicated that diffusivity of wastewater had the greatest effect on dilution factors and that neither discharge rates nor local currents had as much effect. During summer conditions of open water, wastewater is likely to be constrained in a narrow plume close to the coast. Concentrations of sewage bacteria were high around the outfall and detected up to 1.5 km away, along with dye. There were significant differences in sediment concentrations of metals, PBDEs, hydrocarbons, nutrients and faecal sterols between sites within 2 km of the outfall and control sites. We conclude that dilution and dispersal conditions at the Davis outfall are insufficient to prevent the accumulation of contaminants in local sediments and that microbial hazards posed by wastewater are an environmental risk to local wildlife. [ABSTRACT FROM AUTHOR]

Improved testing and remediation procedures for sites contaminated with petroleum hydrocarbons are a priority in remote cold regions such as Antarctica, where costs are higher and remediation times are longer. Isoprenoid/n-alkane ratios are commonly used to determine the extent of biodegradation at low levels but are not useful once the n-alkanes have been removed. This study demonstrates how the diastereomers of the acyclic isoprenoids can be used to determine the extent of biodegradation in moderately biodegraded fuel in soils from a bioremediation trial at Casey Station, Antarctica. The biological diastereomers of pristane (meso; RS = SR) are depleted more rapidly during moderate biodegradation than the geological or mature diastereomers (RR and SS), and thus, the ratio of pristane diastereomers can determine the level of biodegradation. The statistical difference among mean diastereomer ratios for samples grouped according to the biodegradation scale and pristane/phytane ratios was highly significant. The ratios of norpristane and phytane diastereomers also change with biodegradation in a similar fashion, and different levels of sensitivity exist for each. Additional benefits are that the method can be performed on conventional gas chromatographs by non-specialist chemists and that the ratios are independent of evaporation and do not necessarily require a non-biodegraded reference (To) sample. This study details a simple alternative method for determining the extent of biodegradation of fuels at moderate levels that can be applied to a wide range of petroleum products. [ABSTRACT FROM AUTHOR]