Introduction Female sexual dysfunction (FSD) is a multifactorial entity and an underestimated problem with an overall prevalence between 20% and 50%. Prevalence can change according to the diagnostic criteria used, data collection, and others such as psychological and cultural factors. Population studies of FSD are limited, both in Colombia and in other countries. Nevertheless, because of the prevalence found in other studies, FSD can be considered a public health issue that affects the quality of life of women who suffer from it and their partners. Aim To determine the prevalence of FSD and its correlation with the hormonal profile in female outpatients and female staff at San Jose Hospital (Bogotá, Colombia). Methods We performed a cross-sectional study with endocrine measurements. Main Outcome Measures The prevalence of FSD according to the Female Sexual Function Index (FSFI) cutoff points, and androgenic deficiency defined as women with a proportion of free testosterone below 2%. Demographic characteristics, medical history survey, FSFI, and a hormonal profile that included follicle stimulating hormore (FSH), luteinizing hormone (LH), prolactin, estrone, estradiol, sulphate and total dehydroepiandrosterone, androstenedione, sex hormone binding globulin, and total testosterone. Results Fifty (49.5%; 95% confidence interval [CI]: 39.4–59.6%) out of 101 participants met the criteria for FSD; in those women with sexual activity in the past 4 weeks, the prevalence of FSD was 37.8% (95% CI: 27.3–49.1%). In the hormone evaluation sample, 29 out of 32 women with sexual activity in the past 4 weeks met the criteria for FSD. Our results suggest that there is independence between the FSFI scores and the proportion of free testosterone. Conclusions Our findings suggest that FSD in our population has a prevalence as high as that previously reported in the literature; however, because of the sample selection criteria, we could not say that our study results may well reflect the prevalence in our general Colombian population. Garcia S, Moreno S, and Aponte H. Prevalence of sexual dysfunction in female outpatients and personnel at a Colombian hospital: Correlation with hormonal profile.