Objective: To investigate the effect of Vitamin D (VD) combined with Azithromycin (AZM) sequential therapy in Mycoplasma pneumoniae pneumonia (MPP) in children. Methods: 94 patients with MPP admitted to our hospital from January to December 2024 were selected as the study objects. The children were divided into observation group and control group by random number table method, with 61 cases in each group. The control group received AZM sequential therapy; Observation group was additionally treated with VD on the basis of control group. Treatment effectiveness, time to resolve major symptoms, inflammatory factors [Tumor Necrosis Factor-alpha (TNF-alpha), C-reactive protein (CRP)], and adverse drug reactions in the two groups were analyzed and compared. Results: The total effective rate of observation group was significantly higher than that of control group (P < 0.05). The cough relief time, fever remission time and lung rales disappearance time in observation group were significantly shorter than those in control group (P < 0.05). Before treatment, there were no significant differences in CRP and TNF-α levels between the two groups (P > 0.05). After 7 days of treatment, the levels of CRP and TNF-α in both groups were significantly lower than before treatment, and the levels of CRP and TNF-α in observation group were significantly lower than those in control group (P < 0.05). There was no significant difference in the incidence of adverse reactions between the two groups (P > 0.05). Conclusion: Compared with AZM alone, VD-assisted AZM sequential therapy in the treatment of MPP in children can improve the overall therapeutic effect, reduce the inflammatory response of children, promote the relief of symptoms, and do not increase adverse reactions, and has good safety.