Modulation of growth, oxidative stress damage, and trigonelline accumulation in drought-stressed fenugreek by nitrogen-containing biostimulants

Fenugreek (Trigonella foenum-graecum L.) is a medicinal and aromatic crop valued for its diverse bioactive compounds, particularly trigonelline, with wide-ranging applications in pharmaceuticals, food, and cosmetics. As drought stress increasingly threatens global crop productivity and quality, identifying sustainable agronomic interventions is imperative. This study is the first to investigate the combined application of aspartic acid, glutamic acid, and nicotinic acid under drought stress in fenugreek. This work evaluated the impact of varying drought intensities (100%, 80%, and 60% field capacity) combined with foliar applications of nitrogen- containing biostimulants—aspartic acid, glutamic acid, nicotinic acid, and their combined application—on fenugreek growth, antioxidant defenses, chlorophyll fluorescence, membrane integrity, and secondary metabo lite accumulation. Results demonstrated that drought stress diminished shoot biomass, leaf relative water con tent, and total nitrogen, while provoking oxidative stress marked by elevated electrolyte leakage, malondialdehyde accumulation, and altered chlorophyll fluorescence parameters (increased F₀ and F₀/F decreased Fᵥ/F₀ and Fᵥ/F m m ; ). Conversely, drought intensified enzymatic and non-enzymatic antioxidant activities and stimulated trigonelline biosynthesis, indicative of adaptive metabolic responses. Foliar application of bio stimulants significantly alleviated drought-induced damage, with the combined treatment showing superior efficacy in enhancing biomass, water status, proline accumulation, nitrogen content, antioxidant enzyme ac tivities (peroxidase, catalase, ascorbate peroxidase, superoxide dismutase), phenolic compounds, flavonoids, and trigonelline concentration. Individually, aspartic acid and glutamic acid predominantly improved growth and physiological metrics, while nicotinic acid specifically augmented trigonelline levels. This work underscores the potential of targeted amino acid and vitamin-based biostimulants to enhance drought resilience and bioactive compound production in fenugreek, offering a promising strategy for sustainable agriculture and value-added crop development.