Cost and environmental impacts are the two key issues related to the development of biofuels. The size reduction process of feedstock for the production of lignocellulosic ethanol, a second generation biofuel, is energy-intensive and costly. In this study, life cycle analyses and techno-economic analyses were conducted for six different scenarios of corn stover size reduction. Six grinding condition combinations of three different moisture content (5%, 10%, 20%, wet basis) and two different screens (2 mm, 6 mm) of a knife mill were each used for the six scenarios. The cost and environment impact was compared for each scenario on the basis of producing the same amount of fermentable sugars. A comparably cost-efficient and environmentally friendly corn stover size reduction scenario was concluded.