To explore the impact of combined carbon–nitrogen fertilization on the concentrations of Fe (ferrum) and Zn (zinc) in Dendrobium officinale (D. officinale), and to elucidate the underlying metabolic regulatory mechanisms, two-year-old seedlings of D. officinale were selected as the experimental subjects. Three treatment groups were established: a control group (CK), an α-ketoglutaric acid (AKG) treatment group (C treatment, CT), a urea treatment group (N treatment, NT), and an AKG and urea combined treatment group (CT_NT). Samples were collected at 0, 8, 16, 24, and 32 days post-treatment. Physiological and biochemical analyses measured stem contents of iron, zinc, copper, nitrate nitrogen, soluble proteins, and citric acid. Transcriptomic and metabolomic technologies were employed to elucidate molecular mechanisms. Physiological studies have shown that combined carbon–nitrogen application exerts time-dependent regulation on Fe, Zn, and their key metabolites in the stems of D. officinale, presenting a trend of first increasing and then decreasing. Metabolomic analysis revealed that flavonoids, phenolic compounds, and organic acids are involved in Fe chelation, while quercetin, dopamine, and other substances promote Zn absorption. Transcriptomic analysis indicated that combined carbon–nitrogen application activates the accumulation of Fe and Zn contents by upregulating the expression of related genes. Integrated analysis demonstrated that carbon–nitrogen metabolism affects the metabolic network of D. officinale by regulating primary and secondary metabolic pathways. This study elucidated the physiological and molecular mechanisms underlying the regulation of Fe and Zn contents in D. officinale by combined carbon–nitrogen application, providing theoretical support and a scientific basis for the high-efficiency cultivation and quality improvement of D. officinale.

Chinese hickory (Carya cathayensis Sarg.) is a significant woody oil tree species indigenous to China; nevertheless, excessive nitrogen (N) application has emerged as a primary limiting factor for sustainable production, as it induces leaf damage, metabolic imbalance, and premature senescence. α-Ketoglutarate (α-KG), an intermediate of the tricarboxylic acid (TCA) cycle, plays a crucial role in coordinating carbon (C) and N metabolism by providing carbon skeletons for N assimilation. In this study, one-year-old C. cathayensis seedlings were subjected to the following treatments: water control (CK), α-KG alone (40 mg/L, KG3), high urea (0.8%, U), and sequential applications of urea combined with 10, 20, or 40 mg/L α-KG (UKG1–UKG3). Photosynthetic characteristics, antioxidant responses, C–N metabolic parameters, and mineral element contents were evaluated at 0, 4, 8, and 12 weeks after treatment. High-concentration urea application led to visible leaf scorching and curling. At week 8, the urea-only treatment showed no significant difference from the water control in the six OJIP parameters; however, exogenous α-KG application significantly influenced the PSII fluorescence parameters of urea-treated seedlings. Compared with U, UKG3 markedly enhanced PSII performance, with the PI_ABS value rising to 0.614 ± 0.093 (approximately 6.4 times that of U). Although urea treatment increased chlorophy ll content and net photosynthetic rate (Pn), these responses were not accompanied by a significant improvement in PSII photochemical efficiency relative to CK, indicating an imbalance between carbon assimilation and photosynthetic electron utilization under high urea supply. α-KG supplementation also improved nitrogen assimilation-related enzyme activities. GS activity under UKG treatments reached a level comparable to that of KG3. NR activity under UKG treatments decreased compared with both U and KG3 but remained higher than that of CK. α-KG alone enhanced photosynthetic performance, carbon accumulation, GS/Fd-GOGAT activities, and leaf Fe concentration without causing visible stress symptoms. Overall, exogenous α-KG supplementation partially alleviated high-urea-induced physiological disturbances in C. cathayensis seedlings, which was associated with improved PSII function and a more balanced regulation of C–N metabolism. These findings provide physiological evidence supporting the potential application of α-KG in optimizing nitrogen management strategies for C. cathayensis cultivation.