ABSTRACT: The current study had two aims. The primary purpose was to examine the association between serum vitamin D levels and the ergometric evaluation of muscle strength, aerobic capacity, and speed in professional soccer players. The secondary aim was to evaluate the effects of the soccer off-season period on serum vitamin D levels.Sixty-seven Caucasian male soccer players (age 25.6 ± 6.2 and height 1.81 ± 0.08 m), members of two Greek Superleague Soccer teams and one Football-league championship team participated in this study. Exercise performance testing for the determination of squat jump (SJ), countermovement jump (CMJ), 10 (10 m) and 20 meters (20 m) sprint performance, maximal oxygen consumption (VO2max), anthropometry, and blood sampling were performed before (pre) and after (post) the six-week off-season period.Analysis of our results showed the following: (a) a significant correlations between serum vitamin D levels and performance parameters in both pre (SJ; P < 0.001, CMJ; P < 0.001, VO2max; P < 0.001, 10 m; P < 0.001, and 20 m; P < 0.001) and post (SJ; P < 0.001, CMJ; P<0.001, VO2max; P = 0.006, 10 m; P < 0.001, and 20 m; P < 0.001) experimental sessions. (b) Vitamin D concentration increased significantly (P < 0.001) following the six-week off-season period compared to baseline, while at the same time all measured performance parameters decreased (SJ; P < 0.001, CMJ; P < 0.001, 10 m; P < 0.001, 20 m; P < 0.001, VO2max; P<0.001).Our findings suggest that vitamin D levels are associated with the ergometric evaluation of muscle strength, as expressed by SJ and CMJ, sprinting capacity, and VO2max in professional soccer players, irrespective the levels of performance. Furthermore, our data reaffirm the importance of UVB on serum vitamin D levels. Moreover, reductions in exercise training stress may also have beneficial effects on vitamin D levels, suggesting a possible association of its levels and the training-induced stress. Our results indicate a possibly bidirectional interaction between soccer performance indices and vitamin D levels.