In addition to adipocytokines, estradiol (E2) and vitamin D have been reported to affect insulin sensitivity, glucose homeostasis and body weight. However, studies about the impact of E2 and vitamin D on metabolic syndrome (MetS) are still limited. The aim of this study is to clarify the roles of circulating E2 and vitamin D on the risk of MetS in middle-aged Taiwanese males. A total of 655 male volunteers, including 243 subjects with MetS (mean age: 56.7±5.8 years) and 412 normal controls (mean age: 55.1±3.6 years), were evaluated. Subjects with MetS had significantly lower circulating E2, 1,25(OH)2D3, and adiponectin, and higher leptin than those without MetS (P<0.001 for all comparisons). E2 and 1,25(OH)2D3 were significantly associated with 4 individual components of MetS; more than adiponectin and leptin that were only associated with 3 individual components. In multivariate regression analysis, E2 (beta = −0.216, P<0.001) and 1,25(OH)2D3 (beta = 0.067, P = 0.045) were still significant predictors of MetS independent of adiponectin and leptin. Further large studies are needed to confirm our preliminary results and elucidate the possible mechanism.