Innovative Hydroponic Culture of Alkanna tinctoria (L.) Tausch: An Approach Towards Sustainable Extraction Process from Plant Roots

<i>Alkanna tinctoria</i> (L.) Tausch is a valuable medicinal plant known for its root-derived hydroxynaphthoquinone enantiomers, alkannin/shikonin (A/S), which exhibit significant pharmaceutical and cosmeceutical potential. However, its limited natural distribution and overharvesting pose conservation challenges, necessitating sustainable cultivation and extraction strategies. The application of Natural Deep Eutectic Solvents (NaDESs) has garnered significant attention as sustainable alternatives to conventional solvents. However, their toxicity in living plant systems remains largely unexplored. This study presents the successful establishment of an ex situ hydroponic cultivation system using the nutrient film technique (NFT) to grow <i>A. tinctoria</i> under greenhouse conditions. The system promoted plant acclimatization, vigorous root development, and initial production of A/S derivatives. In parallel, the toxicity evaluation of a bio-based NaDES, LeG_5_20 (levulinic acid-glucose, 5:1, with 20% water), applied as a circulating medium, was assessed. Physiological stress responses of the plants to NaDES circulation were assessed through non-destructive measurements, including stomatal resistance, photosynthetic and transpiration rates, and sub-stomatal CO₂ concentration. Short-term (24 min) exposure to NaDES showed no significant adverse effects, while longer exposures (4-8 h) induced marked stress symptoms and loss of leaf area. These findings demonstrate the feasibility of integrating green hydroponic systems with eco-friendly extraction solvents and provide a framework for further optimization of plant age, solvent exposure time, and system design to enable sustainable metabolite recovery without plant destruction.