Chemical profiling and identification of bioactive compounds from biostimulant enhancing salinity tolerance in rice

Panchagavya (PG), a traditional bioorganic formulation, is widely recognized as a biostimulant, enhances growth and development of plants, soil fertility, nutrient uptake and mitigates the different environmental cues. Despite its extensive agricultural and veterinary applications, the precise composition and bioactive compounds responsible for its benefits remain unidentified. While some studies have attempted to isolate its components, standardized extraction and identification techniques are lacking, limiting its targeted application. To address this, we conducted a comprehensive study to isolate and identify compounds from PG. Using column chromatography, four pure compounds were successfully isolated and characterized through electrospray ionization mass spectroscopy (ESI-MS) and Nuclear Magnetic Resonance (NMR) analysis. In silico modelling and molecular docking studies were conducted with eight rice autophagy proteins to explore the molecular interactions, revealing strong binding affinities for all four compounds. Among these, 1-methyl palmitate exhibited the most promising interaction and was selected for further evaluation in rice. Its biological properties were tested in rice seedlings exposed to saline stress through a comprehensive analysis of biochemical, cellular, and molecular approaches. The findings demonstrated that 1-methyl palmitate enhances saline stress tolerance by positively regulating autophagy and programmed cell death pathways, critical for maintaining cellular balance under stress. This regulation was accompanied by significant upregulation of antioxidant genes, autophagy related genes and transcription factors, contributing to the mitigation of salinity. The findings reveal the role of PG as biostimulant in enhancing stress tolerance, optimizing fertilizer efficiency, and contributing to sustainable food production.