The aim of this study was to compare whole and segmental body composition and bone mineral density of collegiate American football players by playing positions. Forty collegiate American football players voluntarily participated in this study. Participants were categorized by playing positions into one of five categories i.e., defensive linemen, offensive linemen, defensive secondary players, offensive secondary players and receivers. Whole body composition and bone mineral density were measured by dual x-ray absorptiometry. Offensive and defensive linemen had higher body mass, a body mass index, lean mass and a fat mass index compared to the remaining three positions and a higher lean mass index compared to offensive secondary players and receivers. Offensive linemen had a higher body fat percentage and lower values of upper to lower lean mass than offensive and defensive secondary players and receivers, and higher total mass to the lean mass ratio and fat mass to the lean mass ratio compared to the other players. Offensive linemen had a higher fat mass index and fat mass to the lean mass ratio than defensive linemen. However, in all other measures they were similar. Offensive and defensive secondary players and receivers were similar with respect to the measured variables. Bone mineral density of the players was within the normal range and no difference in lean mass was observed between the legs. In conclusion, findings of this study showed that the total and segmental body composition profile of collegiate American football players reflected the demands of particular playing positions.