Objectives: Vitamin D is important for the muscular tone and bone metabolism. Due to the high prevalence of vitamin D deficiency in Iranian women, this study aimed to investigate the difference in vitamin D levels between patients with and without pelvic floor disorder (PFD) to assess the possible effect of vitamin D on PFD. Materials and Methods: The present case-control study examined 209 women referring to Ayatollah Rouhani hospital of Babol in 2017. Patients were explored for prolapse and those with at least one disorder or without PFD were categorized as the case (n = 104) or control (n = 105) groups, respectively. Then, the serum level of 25-hydroxy vitamin D was compared between the groups. Results: The mean age and abdominal circumference of the patients were significantly higher in the case group (P < 0.001 & P = 0.046, respectively) as compared to the control group. Meanwhile, the total mean ± SD serum level of vitamin D was 20.03±17.88 ng/ mL and significantly higher in the case group (24.58 ± 20.75 ng/mL) than that of the control group (15.53±13.11 ng/mL), especially in patients with stress and urgency urinary incontinence (SIU/UIU) (both with a significance level of P < 0.001). Conclusions: As a result of PFD, the group with UI, especially the one with SIU or UIU had the highest vitamin D level compared to the control and other groups. Nonetheless, the mean age and educational level were significantly higher and lower in this group, respectively. These results could be due to the multifactorial nature of vitamin D level, that is, it varies based on nutrition, place of residence, and other factors.