Despite a rapid worldwide expansion of the biofuel industry, there is a lack of consensus within the scientific community about the potential of biofuels to reduce reliance on petroleum and decrease greenhouse gas (GHG) emissions. Although life cycle assessment provides a means to quantify these potential benefits and environmental impacts, existing methods limit direct comparison within and between different biofuel systems because of inconsistencies in performance metrics, system boundaries, and underlying parameter values. There is a critical need for standardized life-cycle methods, metrics, and tools to evaluate biofuel systems based on performance of feedstock production and biofuel conversion at regional or national scales, as well as for estimating the net GHG mitigation of an individual biofuel production system to accommodate impending GHG-intensity regulations and GHG emissions trading. Predicting the performance of emerging biofuel systems (e.g., switchgrass cellulosic ethanol) poses additional challenges for life cycle assessment due to lack of commercial-scale feedstock production and conversion systems. Continued political support for the biofuel industry will be influenced by public perceptions of the contributions of biofuel systems towards mitigation of GHG emissions and reducing dependence on petroleum for transportation fuels. Standardization of key performance metrics such as GHG emissions mitigation and net energy yield are essential to help inform both public perceptions and public policy.