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Background/Objectives: Skin wrinkles result from a myriad of multifaceted processes involving intrinsic and extrinsic aging. To combat this effect, plant stem cells offer a renewable and eco-friendly source for various industries, including cosmeceuticals. Salvia miltiorrhiza (SM), which contains the bioactive compound Rosmarinic acid (RA) and has been proposed for its anti-wrinkle effect. Methods: In the present study, calli from SM were cultured and Quality by Design (QbD) was implemented to investigate the effect of different types and concentrations of elicitors; jasmonic acid (JA) and salicylic acid (SA). Both raised RA levels yet, jasmonic acid (50 µM) has resulted in the highest yield for RA, at 16 mg/g. A nanofiber patch was prepared and characterized in-vitro by the release percentage, drug content, swelling degree, scanning electron microscope, and surface roughness. Then, the anti-wrinkle effect of the patch was tested in a UV wrinkle-induced mouse model. Results: Interestingly, after treatment, there were visibly fewer wrinkles, and the skin was softer than in the untreated control group. This study suggests that the treatment exerted its effect through the Nrf2/Keap1 pathway, which plays a crucial role in cellular antioxidant protective processes. By activating this pathway through boosting Nrf2 and diminishing Keap1 cellular content, the nanofiber patch enhances the production of antioxidant enzymes, such as superoxide dismutase and glutathione peroxidase, enhancesglutathione, and reduces the skin lipid peroxidation, collectively indicating enhanced skin quality. Conclusions: In conclusion, this study highlights the importance of this formula as an anti-wrinkle treatment, and future clinical studies are recommended to further unveil the potential of this formula. [ABSTRACT FROM AUTHOR]