Background/Objectives: This study evaluated the associations of shift work with serum vitamin D levels, excessive daytime sleepiness (EDS), and symptoms of anxiety and depression among female nurses working at a university hospital. Methods: This was an analytical cross-sectional study. Sociodemographic and occupational characteristics were collected via a structured questionnaire. EDS was assessed using the Epworth Sleepiness Scale (ESS), and symptoms of anxiety and depression were assessed using the Hospital Anxiety and Depression Scale (HADS). Serum 25-hydroxyvitamin D [25(OH)D] levels were measured by electrochemiluminescence (Elecsys Vitamin D total II, cobas e 801). Deficiency was defined as <20 ng/mL. Results: Mean age was 32.4 ± 6.8 years; 44.9% worked night shifts. Mean 25(OH)D level was 13.16 ± 7.87 ng/mL, with deficiency in 84.3% and insufficiency in 13.5%. EDS was present in 30.8%, anxiety symptoms in 26.9%, and depressive symptoms in 46.2%. Vitamin D levels were lower in night-shift nurses, though not significantly (p = 0.056). In univariable logistic regression analysis, night-shift work was associated with vitamin D deficiency (OR = 2.053; 95% CI: 1.068–3.948; p = 0.031), but this association was not significant after adjustment (aOR = 1.136; 95% CI: 0.541–2.389; p = 0.736). EDS was associated with anxiety symptoms (aOR = 2.795; 95% CI: 1.553–5.031; p = 0.001) and depressive symptoms (aOR = 1.887; 95% CI: 1.077–3.306; p = 0.027). Anxiety and depressive symptoms were associated with each other (aOR = 7.332 and 7.285; p < 0.001). Conclusions: A high prevalence of vitamin D deficiency, EDS, anxiety symptoms and depressive symptoms was observed among female nurses. Vitamin D status was not significantly associated with EDS, anxiety symptoms, or depressive symptoms, whereas EDS was associated with both anxiety and depressive symptoms.
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