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Vitamins are essential for human health. In terms of local application for wound healing, vitamins’ positive effect remains unclear. However, because of the regular appearance of nutritional deficiency in chronic wound patients, a favorable impact of locally applied vitamins can be hypothesized.Vitamins B3, B5, B6, B7, B9, B10, B12, and C individually as well as different combinations of B vitamins were investigated regarding their ability to promote proliferation and migration of human skin fibroblasts and keratinocytes. Proliferation assays with and without bacterial challenge, immunocytochemical staining, and scratch assay were used to determine the most effective combination(s).Some vitamin combinations showed a positive impact on proliferation, especially for keratinocytes after 72 hours. In terms of wound closure, the combinations B9 and B12; B3, B5, B6, and B10; and B3, B5, and B7 improved closure rates by 25% to 30%. The improved closure rates are also reflected by immunocytochemically detected upregulation of the migration marker CXCR4 for several combinations.Certain combinations of B vitamins demonstrate a positive influence on human keratinocytes and fibroblasts. Vitamins especially promoted fibroblast migration, and a statistically significant induction of keratinocyte proliferation was observed. Therefore, local vitamin application could benefit the physiologic wound healing process.