Plant density was varied with P, Ca, Mg, and KNO3 in a multifactor experiment to improve Curcuma longa L. micropropagation, biomass and microrhizome development in fed-batch liquid culture. The experiment had two paired D-optimal designs, testing sucrose fed-batch and nutrient sucrose fed-batch techniques. When sucrose became depleted, volume was restored to 5% m/v sucrose in 200 ml of modified liquid MS medium by adding sucrose solutions. Similarly, nutrient sucrose fed-batch was restored to set points with double concentration of treatments’ macronutrient and MS micronutrient solutions, along with sucrose solutions. Changes in the amounts of water and sucrose supplementations were driven by the interaction of P and KNO3 concentrations. Increasing P from 1.25 to 6.25 mM increased both multiplication and biomass. The multiplication ratio was greatest in the nutrient sucrose fed-batch technique with the highest level of P, 6 buds/vessel, and the lowest level of Ca and KNO3. The highest density (18 buds/vessel) produced the highest fresh biomass at the highest concentrations of KNO3 and P with nutrient sucrose fed-batch, and moderate Ca and Mg concentrations. However, maximal rhizome dry biomass required highest P, sucrose fed-batch, and a moderate plant density. Different media formulations and fed-batch techniques were identified to maximize the propagation and storage organ responses. A single experimental design was used to optimize these dual purposes.

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BackgroundTurmeric is a rich source of bioactive compounds useful in both medicine and cuisine. Mineral concentrations effects (PO43−, Ca2+, Mg2+, and KNO3) were tested during in vitro rhizome development on the ex vitro content of volatile constituents in rhizomes after 6 months in the greenhouse. A response surface method (D-optimal criteria) was repeated in both high and low-input fertilizer treatments. Control plants were grown on Murashige and Skoog (MS) medium, acclimatized in the greenhouse and grown in the field. The volatile constituents were investigated by GC-MS.ResultsThe total content of volatiles was affected by fertilizer treatments, and in vitro treatment with Ca2+ and KNO3; but PO43− and Mg2+ had no significant effect. The content was higher in the high-input fertilizer treatments (49.7 ± 9 mg/g DM) with 4 mM Ca2+, 60 mM KNO3 and 5 mM NH4+, than the low-input fertilizer (26.6 ± 9 mg/g DM), and the MS control (15.28 ± 2.7 mg/g DM; 3 mM Ca2+, 20 mM K+, 39 mM NO3−, 20 mM NH4+, 1.25 mM PO43−, and 1.5 mM Mg2+). The interaction of Ca2+ with KNO3 affected curcumenol isomer I and II, germacrone, isocurcumenol, and β-elemenone content. Increasing in vitro phosphate concentration to 6.25 mM increased ex vitro neocurdione and methenolone contents.ConclusionThese results show that minerals in the in vitro bioreactor medium during rhizome development affected biosynthesis of turmeric volatile components after transfer to the greenhouse six months later. The multi-factor design identified 1) nutrient regulation of specific components within unique phytochemical profile for Curcuma longa L. clone 35–1 and 2) the varied phytochemical profiles were maintained with integrity during the greenhouse growth in high fertility conditions.Electronic supplementary materialThe online version of this article (10.1186/s12870-018-1345-y) contains supplementary material, which is available to authorized users.

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