DDGS could have higher market price and wider use if it could be separated into higher protein and higher fiber fractions. In our work, DDGS was firstly sieved into three size categories, and one category was further separated into light, midlight, midheavy and heavy fractions using a gravity separator. This process was effective in getting enhanced DDGS with increased protein and oil. In this study, both Life Cycle Assessment (LCA) and Techno-Economic Analysis (TEA) of our approach to DDGS fractionation were conducted. Three scales, including lab scale, pilot scale, and commercial scale of DDGS fractionation were considered and analyzed. All equipment parameters were obtained from industrial manufacturers. Both the environmental impact and the cost per unit of DDGS fractionation decreased as the fractionation scale expanded.