New biological trends on cell and callus growth and azadirachtin production in Azadirachta indica

Azadirachtin is an important secondary metabolite from Azadirachta indica used as a natural biopesticide. This study is the first comprehensive report concerning the influence of plant growth regulators on callus induction, cell suspension growth, and azadirachtin accumulation and production in cell suspension cultures of A. indica. We investigated the effect of plant growth regulators including different types of auxins and cytokinins and their combinations on callus induction, cell suspension growth, and azadirachtin accumulation and production. The highest percentage of callusing (100%) obtained at different combinations of plant growth regulators on MS medium supplemented with 1 mg/L picloram and 2 mg/L kinetin and the highest fresh weight of callus (264.50 mg) was observed in MS medium containing 1.5 mg/L NAA and 3 mg/L kinetin. In cell suspension cultures, the maximum cell density, SCV, and PCV were 2.44 × 106 cells per mL, 97.95%, and 81.46%, respectively, obtained in the MS medium containing 1.5 mg/L 2,4-D and 3 mg/L zeatin riboside. The highest average growth rate (0.25 days) was on MS medium containing 1.5 mg/L NAA and 3 mg/L zeatin riboside. The MS medium supplemented with 1 mg/L picloram and 2 mg/L kinetin produced the highest amount of fresh cell weight (493.02 g/L), dry cell weight (77.27 g/L), azadirachtin accumulation (3.69 mg/gDW), and azadirachtin production (285.64 mg/L). The results showed that all measured indices had positive correlation with together except FCW and DCW with azadirachtin accumulation.