Background: Vitamin D plays a critical role in bone metabolism and mineralization, which are essential for optimal fracture healing. Deficiency in serum vitamin D may delay the bone healing process, particularly in long bone fractures like the tibia. This study aimed to evaluate serum vitamin D levels in patients with tibial shaft fractures and assess their correlation with bone healing time. Materials and Methods: A longitudinal observational study was conducted on 70 adult patients (aged 20–60 years) with closed tibial shaft fractures treated conservatively or surgically. Serum 25-hydroxyvitamin D levels were measured at admission. Patients were categorized into vitamin D sufficient (≥30 ng/mL), insufficient (20–29 ng/mL), and deficient (<20 ng/mL) groups. Bone healing was assessed clinically and radiographically at monthly intervals until confirmed union. Correlation analysis was performed between vitamin D status and time to union. Results: Out of 70 patients, 28 (40%) were vitamin D deficient, 22 (31.4%) insufficient, and 20 (28.6%) sufficient. The mean healing time was significantly longer in the deficient group (19.2 ± 2.8 weeks) compared to the sufficient group (13.5 ± 2.1 weeks) (p < 0.001). A strong negative correlation was found between serum vitamin D levels and bone healing time (r = -0.62, p < 0.001). Delayed union (>20 weeks) was observed in 32.1% of deficient patients compared to 5% in the sufficient group. Conclusion: Low serum vitamin D levels are significantly associated with prolonged bone healing time in tibial shaft fractures. These findings underscore the importance of screening and correcting vitamin D deficiency in fracture patients to promote timely bone regeneration.