OBJECTIVE: In this study, we isolated the milk sphingolipid-enriched fraction (MSEF) of sweet buttermilk powder and conducted a clinical trial for evaluating its efficacy in skin barrier recovery. METHODS: Milk sphingolipid-enriched fraction was isolated via solvent extraction of buttermilk powder, and further concentrated by removing the phospholipids and neutral lipids. A cream containing 1% MSEF was used during clinical trials to assess for water holding and skin barrier recovery capacities. RESULTS: The main components of the MSEF were sphingomyelin, glucosylceramide and lactosylceramide, confirmed by TLC, HPLC, MS and NMR. The MSEF cream-treated group had a significantly higher (P < 0.05) water holding capacity, compared with the base cream (vehicle) group. Compared with that in the base cream group, transepidermal water loss (TEWL) recovery increased in the presence of the sphingolipid-containing MSEF cream (MSEF group), with a significant difference (P < 0.05) recorded on day 14. CONCLUSION: The MSEF cream contributed to improving the water holding capacity and skin barrier recovery of damaged skin. Therefore, sphingolipid-containing MSEF can be useful for strengthening or repairing skin barrier function. strength, and gene mutations or polymorphisms in ceramide biosynthesis affect skin barrier function [10].