Research was conducted to determine the effect of nutrient management practices on bio-fuel crop production, and to evaluate long term effects of bio-fuel crop production on selected chemical, physical and microbiological properties. Experimental plots for research on bio-fuel crops production were established in 2008 at two sites in Missouri. The experimental design was an 8x3 factorial laid out in a split plot design. The main plots were eight bio-fuel cropping systems including: 1) continuous corn for grain (CCG), 2) continuous corn for grain and stover removal (CCGS), 3) corn-soybean rotation for grain (CSG), 4) soybean-corn rotation for grain (SCG), 5) sweet sorghum-wheat double crop (SSW), 6) miscanthus (MIS), 7) switch grass (SWI) and 8) tall fescue (TF). Subplots received the following fertilizer treatments: 1) University of Missouri recommended P and K with a 4 year buildup; 2) fertilizer recommendations based on annual crop removal values; and 3) control without P and K. Soil samples were collected for soil fertility analysis (pH, P, K, Ca, Mg, OM and CEC), organic and total N, wet aggregate stability, and potential C and N mineralization by measuring dehydogenase (DEHYD) and glucosaminidase (GLUCO) enzyme activities using in vitro assays. Grain yield, dry matter production and nutrient uptake were measured. This presentation reports results of the nutrient uptake and fertility analysis. In continuous corn cropping system P and K applications based upon maintenance and buildup had the highest grain yield; however, stover yield was greatest when P and K application was based upon removal. Luxury P and K consumption and removal were found in tall fescue and sweet sorghum. In the first two years of this study, it is apparent that in order to maximize dry matter and grain yield in a monoculture system P and K levels will need to be closely monitored.