Salycilic Acid Induces Exudation of Crocin and Phenolics in Saffron Suspension-Cultured Cells

The production of crocin, an uncommon and valuable apocarotenoid with strong biologicalactivity, was obtained in a cell suspension culture of saffron (Crocus sativus L.) established fromstyle-derived calli to obtain an in-vitro system for metabolite production. Salycilic acid (SA) wasused at different concentrations to elicit metabolite production, and its effect was analyzed aftera 4 days of treatment. HPLC-DAD analysis was used for total crocin quantification while theFolin-Ciocâlteu method was applied for phenolic compounds (PC) content. Interestingly, despitecell growth inhibition, a considerable exudation was observed when the highest SA concentrationwas applied, leading to a 7-fold enhanced production of crocin and a 4-fold increase of phenolicscompared to mock cells. The maximum antioxidant activity of cell extracts was evidenced after SA0.1 mM elicitation. Water-soluble extracts of saffron cells at concentrations of 1, 0.5, and 0.1 µg mL-1showed significant inhibitory effects on MDA-MB-231 cancer cell viability. The heterologous vacuolarmarkers RFP-SYP51, GFPgl133Chi, and AleuRFP, were transiently expressed in protoplasts derivedfrom the saffron cell suspensions, revealing that SA application caused a rapid stress effect, leadingto cell death. Cell suspension elicitation with SA on the 7th day of the cell growth cycle and 24 hharvest time was optimized to exploit these cells for the highest increase of metabolite production insaffron cells.