To investigate circulating hormonal, metabolic and inflammatory biomarker profiles in obese and non-obese middle-aged women. Methods A total of 110 women, aged 40-60 years, were included in this cross-sectional study. Patients were allocated, according to the occurrence of menopause and body mass index (BMI), into four groups: PM0 (premenopausal non-obese), PM1 (premenopausal obese), M0 (postmenopausal non-obese), and M1 (postmenopausal obese). Serum levels of gonadotropins, sex hormones, lipid markers, leptin, hs-CRP and interleukin-6 were obtained using either colorimetric or immunoenzymatic assays. Univariate and correlation analyses were performed among all clinical and laboratorial parameters. Principal component analysis was used to characterize subsets of biomarkers, which had their discriminatory capacity tested using discriminant function analysis. Results Levels of gonadotropins and female sex hormones were similar between PM0 and PM1 and between M0 and M1 (p > 0.05), all of them varied between PM0 and M0 (p < 0.05), but only estradiol was significantly altered in the comparison between PM1 and M1 (p = 0.027). Regarding metabolic markers, leptin was lower in PM0 than in M0 (p = 0.010) and higher in M1 than in M0 (p = 0.046). In premenopausal women, BMI correlated only to leptin, while it correlated to several other markers in postmenopausal women. A combination of FSH and leptin serum levels significantly discriminated the four groups (Wilks's lambda < 0.001, in canonical functions 1 and 2).