The aim of the study was to compare the players characteristics and physical fitness parameters according to playing positions and to identify the relation between body composition and physical fitness in professional soccer players at the beginning of the preparation period. Sixteen professional soccer players participated in this study. Body composition was evaluated by air displacement plethysmography instrumentation (ROD POD (R)) and physical fitness with specific tests: Running Anaerobic Sprint Test (RAST), Counter Moviment Jump (CMJ), Squat Jump (SJ) and Abalakov jump (ABK),Yo-yo Intermittent Recovery Leven(Yo-yo IR1), agility test and flexibility. Kruskal Wallis test demonstrated no significant difference in all estimated body composition, age, height and physical fitness parameters according to three playing positions (p < 0.05). We verified a significant negative correlation between body fat percentage and the performance of CMJ, SJ, ABK, peak, mean and minimum power (-.51 to -.87) and a significant positive correlation with the agility parameter (r = .85). Regarding to lean mass percentage, a significant positive correlation was found with performance of CMJ, SJ, ABK and peak, mean and minimum power (.51 to - .82) and significant negative correlation with agility parameter (r = -.85). Based on the results, we concluded that at the beginning of a pre-season a higher body fat percentage is associated with a negative influence in physical fitness, thus confirming that a greater amount of lean mass percentage is positively associated with the initial level of physical fitness of professional soccer players.