The Antioxidant and Tyrosinase-Inhibiting Activities of 4-Hydroxycinnamoyl-Malate from a Citrus junos Callus Extract.

We evaluated the antioxidant and skin-lightening activities of 4-hydroxycinnamoyl-malate (4-HCM) in vitro ; the material is a water-soluble component of Citrus junos callus. 4-HCM was waterextracted from callus grown on MS medium containing picloram to enhance growth. The antioxidant activity of 4-HCM was determined using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis-(3-ethylbenzoline-6-sulfonate) (ABTS) free-radical-scavenging assays. 4-HCM-mediated inhibition of extracellular matrix protein glycation was assessed using the elastin and collagen glycation assays. Inhibition of skin pigmentation was evaluated by measuring anti-tyrosinase activity and melanogenesis in melanoma cells. 4-HCM was then incorporated into elastic nanoliposomes (ENLs) and delivered topically to enhance percutaneous absorption. At 5 mM, the free-radical-scavenging activity of 4-HCM was 54.2±0.631% in the ABTS assay, comparable to that of 1 mM ascorbic acid (46.5± 0.17%). The IC ₅₀ value for inhibition of advanced glycation end-product formation from elastin was 1.25±0.23 mM; collagen glycation was completely inhibited when the 4-HCM level was >5 mM. At 0.5 mM, the material afforded 49.2±3.09% inhibition of tyrosinase activity and, at 10 mM, reduced the melanocyte melanin content by 78% without significant cellular toxicity. Moreover, 4-HCM-loaded ENLs exhibited 569% enhanced permeation of 4-HCM across the human epidermis, which may afford skin-lightening if included in cosmetic formulations.