Lamiophlomis rotata (Benth.) Kudo, an endangered medicinal plant endemic to the Qinghai-Tibet Plateau (altitude of 3100–5000 m), faces conservation challenges due to wild resource scarcity and cultivation difficulties. This investigation employed a gradient nitrogen stress system (N1-N4) with a nitrogen-free control (CK) to decipher its nitrogen adaptation mechanisms. Key findings revealed: (1) superior biomass accumulation, SPAD content, and total nitrogen levels in N1 compared to N4; (2) N4-induced suppression of antioxidant enzymes concurrent with elevated secondary metabolites; (3) transcriptomic perturbations in nitrogen metabolism, monoterpenoid biosynthesis, and photosynthetic pathways; (4) identification of eight nitrogen metabolism-related genes and eight iridoid synthase-associated genes exhibiting stress intensity-dependent expression patterns. Notably, key enzymatic activities (NR, Nrt, CA, GS, FM, DXS, NM) demonstrated unimodal responses peaking at N3 intensity. The species demonstrated nitrogen conservation strategies through enhanced assimilation and metabolic diversion into secondary biosynthesis, providing critical insights for alpine medicinal plant cultivation.

Gentiana lawrencei var. farreri (G. farreri) is an important endangered Tibetan medicinal plant. This study aimed to uncover temperature-mediated physiological and molecular mechanisms underlying growth and flowering of this endangered Tibetan medicinal herb, providing theoretical guidance for its standardized artificial cultivation. This study established three diurnal temperature treatments: 15/5 °C (S15), 20/10 °C (S20), and 25/15 °C (S25), and conducted integrated analyses of phenotypic, physiological, non-targeted metabolomic, and transcriptomic data. The results showed that S15 significantly promoted plant biomass accumulation and flowering, with a flowering rate of 45.83%, significantly higher than that of other treatments. S15 significantly reduced the levels of soluble sugar, soluble protein, fructose, and starch, while increasing CAT activity to enhance antioxidant capacity; endogenous hormones showed elevated gibberellin (GA) and decreased indole-3-acetic acid (IAA), establishing a hormonal balance favorable for flowering. Integrated metabolomic and transcriptomic analysis revealed that the flavonoid biosynthesis pathway (ko00941) is the core pathway responsive to temperature; S15 induces an increase in the contents of flavonoid metabolites including naringenin, delphinidin, petunidin, and peonidin, and simultaneously upregulates the expression of key genes including the PAL-encoding gene Cluster-62424.2, the 4CL-encoding gene Cluster-45063.0, and the CHS-encoding gene Cluster-44799.3 involved in flavonoid biosynthesis under this condition. S15 reduces the content of osmotic adjustment substances, increases GA content in plant branches, and enhances antioxidant capacity, which synergistically promotes the growth and flowering of G. farreri . Additionally, S15 mediates the accumulation of flavonoid compounds by regulating key genes involved in flavonoid biosynthesis, thereby elevating the accumulation of bioactive constituents isoorientin and isoscoparin-2″-O-β-D-glucopyranoside. This study can provide crucial theoretical support for the optimal temperature regulation and control in the artificial cultivation of G. farreri.