Exogenous NO Induces ROS-Mediated Physiological Stress and Enhances Codonopsis pilosula Quality

To investigate the effects of exogenous nitric oxide (NO) on secondary metabolism and concomitant quality formation of the medicinal herb Radix Codonopsis, fresh Codonopsis pilosula roots were treated with sodium nitroprusside (SNP, an NO donor) at concentrations of 0, 0.1, 0.5, 2.5, 5.0 and 7.5 mmol/L for five consecutive days. Reactive oxygen species ( $${\text{O}}_{2}^{{\bullet - }}$$ and H2O2), malondialdehyde (MDA), five target secondary metabolites, and 1,3-diphosphoglycerate (1,3-DPG) were measured. Activities of antioxidant enzymes (SOD, CAT, POD) and secondary-metabolism-related enzymes (PAL, Δ12-FAD2, MVD, FDFT1) were quantified. SNP treatments (0.1–7.5 mmol/L) raised endogenous NO, $${\text{O}}_{2}^{{\bullet - }}$$ , H2O2 and MDA and activated SOD, CAT and POD in C. pilosula, indicating that SNP triggered stress‑related physiological responses. PAL, FAD2 and MVD activities rose significantly from day 1 to day 4, while FDFT1 peaked on day 2, collectively facilitating secondary‑metabolite accumulation. For the 0.1 mmol/L SNP group, tangshenoside I, codonoposide, syringin, atractylenolide II and atractylenolide III increased by 175.4, 211.3, 11.1, 9.4 and 2.9% on day 4, relative to day 0; the control showed slight changes. As a key sugar‑catabolism‑derived biosynthetic precursor, 1,3‑DPG had a 61.8% rise, meaning that SNP treatment facilitated secondary metabolic biosynthesis. Secondary metabolites are widely recognized as active substances in medicinal plants; pharmacological tests showed SNP‑treated Radix Codonopsis improved hematological profiles, secretory immunoglobulin A (sIgA), immunoglobulin G (IgG), spleen and thymus indices (P < 0.05) versus ordinary Radix Codonopsis. In conclusion, SNP induces a stress-adaptive physiological state in C. pilosula, stimulates secondary metabolite biosynthesis, and ultimately improves the medicinal quality of Radix Codonopsis.