The second‐to‐fourth digit ratio (2D:4D) is the putative marker of prenatal hormone exposure. The 2D:4D ratio or the right–left difference (Dr‐l) are said to be negative and positive correlates, respectively, of circulating testosterone and estrogen in both adult males and females. However, previous studies on the subject have reported mixed results. This study aimed to determine the sex‐moderated relationship between the 2D:4D ratio and adult circulating testosterone, estradiol, testosterone‐to‐estradiol ratio and the free androgen index. This was a cross‐sectional study from January to June 2021 at the University for Development Studies, Ghana. The study involved 62 participants (Female = 28; Male = 34), aged between 20 and 26 years. The right (2D:4DR), the left (2D:4DL), and their difference (Dr‐l) were measured by computer‐assisted analysis. Fasting venous samples were assayed for total testosterone (T), estradiol (E2), and sex hormone‐binding globulin (SHBG) using ELISA. The free androgen index (FAI) was then calculated (T/SHBG) and the data were analyzed using moderated and/or weighted regression. Males had significantly higher T and FAI than females while females had significantly higher E2 than males, which were independent of age and body mass index (p < 0.001). There was a significant SEX*Dr‐l interaction on FAI (p = 0.007). The Dr‐l correlated negatively with FAI in males but positively in females and accounted for about 94.0% of the variability of FAI in males (adjR2 = 0.940) and only 0.2% in females (adjR2 = 0.002). The 2D:4D ratio, a putative marker of prenatal hormone exposure, may have an impact on sex differences in adult free androgen index.