Patients with atopic dermatitis(AD) have impaired barrier function, which decreases skin hydration, weakens their defense against microorganisms, and culminates in increased inflammatory responses. Here, we conducted a clinical trial to evaluate the efficacy of a multi-lamellar emulsion (MLE) containing the pseudoceramide PC-9S and a synthetic sphingosine kinase 1 (SPHK1) activator, Defensamide™, in improving mild-to-moderate atopic dermatitis. Forty patients aged ≥ 2years were randomized into a combined-therapy group treated with the MLE containing PC-9S and Defensamide™ plus a topical corticosteroid and a topical-corticosteroid-only group. Assessments based on therapeutic methods included the Eczema Area and Severity Index (EASI), the Investigator Global Assessment (IGA), transepidermal water loss (TEWL), stratum corneum hydration (SCH), skin dryness, a visual analogue scale (VAS) of itchiness, a VAS of sleep disturbance, patientsatisfaction, and the Dermatology Life Quality Index (DLQI). Thirty-eight patients completed this study. In the combined-therapy group, significant improvements in clinical and instrumental measures such as EASI scores, skin hydration, and skin dryness were noted at 4weeks compared to baseline, but such improvements were not noted in the topical corticosteroid-only group. Subjective assessments of itching and sleep disturbance and DLQI scores also showed significant improvements in the combined-therapy group. Combined therapy with the MLE containing Defensamide™ and PC-9S and with topical corticosteroid demonstrated superior clinical outcomes compared with topical corticosteroid monotherapy. Our findings underscore the potential of MLE-containing formulations as effective adjunctive therapies for AD, offering both objective and subjective symptomatic relief and enhancing patients' quality of life.