Influence of light type, cadmium, and glycine concentration on vinblastine and vincristine production in Catharanthus roseus cell suspension culture with response surface methodology-based optimization of elicitor condition

The Catharanthus roseus contains anticancer alkaloids such as vinblastine and vincristine and has potent medicinal properties. In this study, the effects of different concentrations of plant growth regulators on callus induction, various types of LED light, cadmium and glycine levels, and sampling times on cell suspension growth and the synthesis of vinblastine and vincristine were examined. The optimal conditions were predicted using response surface methodology. Additionally, the expression of the DAT and D4H genes was investigated. Results showed that calluses obtained in B5 medium containing 1 mg/L 2,4-D were suitable for establishing and growing suspension cultures. Among different LED lights, cell suspensions grown in the dark exhibited better growth and higher accumulation and production of vinblastine and vincristine. The highest cell biomass was achieved with 50 µM cadmium plus 2 mg/L glycine for 24 h. The greatest accumulation and production of vinblastine (9.41 mg/g DW and 164.73 mg/L) occurred with 100 µM cadmium plus 8 mg/L glycine for 48 h. The highest vincristine accumulation and production (22.73 µg/g DW and 389.99 µg/L) were observed with 50 µM cadmium plus 2 mg/L glycine for 24 h. Predicted optimal conditions for maximum vinblastine accumulation and production were 150 µM cadmium plus 3.6 mg/L glycine for 72 h (9.59 mg/g DW) and 102 µM cadmium plus 6.24 mg/L glycine for 52 h (133.61 mg/L). For vincristine, optimal conditions were predicted at 50 µM cadmium plus 2 mg/L glycine for 51.5 h (21.94 µg/g DW) and 102 µM cadmium plus 6.24 mg/L glycine for 47.8 h (219.26 µg/L). The analysis of DAT and D4H gene expression showed that both genes expression increased significantly across all treatments, with the highest vinblastine accumulation.