Laboratory evaluation of oil spill bioremediation products in salt and freshwater systems

Ten oil spill bioremediation products were tested in the laboratory for their ability to enhance biodegradation of weathered Alaskan North Slope crude oil in both freshwater and saltwater media. The products included nutrients to stimulate inoculated microorganisms, nutrients plus an oil-degrading inoculum, nutrients plus compounds intended to stimulate oil-degrading activity, or other compounds intended to enhance microbial activity. The product tests were undertaken to evaluate significant modifications in the existing official United States Environmental Protection Agency (EPA) protocol used for qualifying commercial bioremediation agents for use in oil spills. The EPA protocol was modified to include defined formulas for the exposure waters (freshwater, saltwater), a positive control using a known inoculum and nutrients, two negative controls (one sterile, the other inoculated but nutrient-limited), and simplified oil chemical analysis. Three analysts conducted the product test independently in each type of exposure water in round-robin fashion. Statistical tests were performed on analyst variability, reproducibility, and repeatability, and the performance of the various products was quantified in both exposure media. Analysis of variance showed that the analyst error at each time-point was highly significant (P values ranged from 0.0001 to 0.008, depending on water type and oil fraction). In the saltwater tests, six products demonstrated various degrees of biodegradative activity against the alkane fraction of the crude oil and three degraded the aromatic hydrocarbons by >10%. In the freshwater tests, eight products caused >20% loss of alkane hydrocarbons, of which five degraded the alkanes by >50%. Only four products were able to degrade polycyclic aromatic hydrocarbons (PAHs) by >20%, one of which caused 88% removal. However, when the variability of the analysts was taken into consideration, only one of the ten products was found to yield significant percent removals of the PAH fraction and only in freshwater. Viable microorganism population analysis (most-probable-number method) was also performed on every sample by each operator to measure the changes in aromatic and alkane hydrocarbon-degrading organism numbers. In general, little evidence of significant growth of either alkane- or PAH-degraders occurred among any of the ten products in either the saltwater or freshwater testing.