A microcosm-based experiment was conducted to investigate the ability of community level physiological profiling (CLPP) to detect changes in an aquatic microbial community resulting from exposure to oil sands process affected water (OSPW). Detection of the microbial response was done by using the Biolog EcoPlate system, a commercially-available system originally developed for the assessment of rhizosphere microbial communities. The Biolog system consists of a 96-well microtitre plate. Each well on the plate contains both a pure organic compound (the substrate), and a tetrazolium dye. When a microorganism metabolizes the substrate, the dye is reduced into a purple formazan product. The purple colour of each well is characterized using a spectrophotometer measuring optical density (OD) at 590 nm. In this study, we used the EcoPlate version of the Biolog System. Reductions in metabolic activity and inoculum density were detected in the high OSPW group. Overall, indicators of microbial metabolic activity decreased over time. One of these indicators, the sum of substrate means (SSM), showed a dramatic response to weekly water changes. Low cyclicity naphthenic acids demonstrated a reduction over the first and last weeks of the exposure period. Higher cyclicity naphthenic acids demonstrated reductions in the first but not the last week of exposure. The total naphthenic acid (TNA) content of the microcosms appeared to increase over the last week of the exposure period, which may reflect the accumulation of products of microbial metabolism. Our results suggest that inoculum density remains a source of variability for CLPP results. Furthermore, the biological context under which the microbial community forms has a strong influence on its metabolic characteristics. The changes in naphthenic acid concentration (total and speciated) likely reflect adsorption and/or microbial metabolism. Our observation of increased phytoplankton in the presence of OSPW is consistent with the available literature. Additional research will be required to determine if this finding can be developed into an indicator of toxic effect, rather than just the presence/concentration of OSPW.