INTRODUCTION AND HYPOTHESIS: Our aim was to characterize the relationship between 25-hydroxyvitamin D [25(OH)D] status with pelvic floor symptom distress and impact on quality of life. METHODS: A retrospective chart review was performed in women with a 25(OH)D level drawn within 1 year of their gynecology/urogynecology visit. Validated questionnaires including the Colorectal-Anal Distress Inventory (CRADI)-8 and Incontinence Impact Questionnaire (IIQ-7) were used. Multivariate analyses characterized pelvic floor disorder (PFD) symptom differences among women by vitamin D status. RESULTS: We studied 394 women. Mean ± standard deviation (SD) 25(OH)D levels were higher in women without than with PFD symptoms (35.0 ± 14.1 and 29.3 ± 11.5 ng/ml, respectively (p < 0.001)]. The prevalence of vitamin D insufficiency was 51% (136/268). CRADI-8 and IIQ-7 scores were higher among women with vitamin D insufficiency (p = 0.03 and p = 0.001, respectively). Higher IIQ-7 scores were independently associated with vitamin D insufficiency (p < 0.001). CONCLUSIONS: Insufficient vitamin D is associated with increased colorectal symptom distress and greater impact of urinary incontinence on quality of life.