Objective: To evaluate the effect of monophasic combined oral contraceptive pill (COCP) and menstrual cycle phase in healthy young women on white matter (WM) organization using diffusion tensor imaging (DTI). Methods: Thirty young women were included in the study; 15 women used COCP and 15 women had a natural cycle. All subjects underwent DTI magnetic resonance imaging during the follicular and luteal phase of their cycle, or in different COCP cycle phases. DTI parameters were obtained in different WM structures by performing diffusion tensor fibre tractography. Fractional anisotropy and mean diffusivity were calculated for different WM structures. Hormonal plasma concentrations were measured in peripheral venous blood samples and correlated with the DTI findings. Results: We found a significant difference in mean diffusivity in the fornix between the COCP and the natural cycle group. Mean diffusivity values in the fornix were negatively correlated with luteinizing hormone and estradiol blood concentrations. Conclusion: An important part in the limbic system, the fornix, regulates emotional processes. Differences in diffusion parameters in the fornix may contribute to behavioural alternations related to COCP use. This finding also suggests that the use of oral contraceptives needs to be taken into account when designing DTI group studies. Key Points: • Diffusion tensor MRI offers new insights into brain white matter microstructure. • The effects of oral hormonal contraception were examined in young women. • Diffusion tensor images and hormone blood concentrations were evaluated. • Women using hormonal contraception demonstrated higher mean diffusivity in the fornix. • These changes may contribute to behavioural alternations related to contraception use.