Anti-Hypoxic Phytochemicals in Gao-Shan-Hong-Jing-Tian Oral Liquid: LC-MS Profiling, Network Pharmacology, and Carbonic Anhydrase Inhibition

Gao-shan-hong-jing-tian (GSHJT) Oral Liquid is a phytochemical-rich preparation derived from Rhodiola, yet its anti-hypoxic active constituents and molecular mechanisms remain poorly understood. This study aimed to identify the key anti-hypoxic phytochemicals in GSHJT Oral Liquid and clarify their mechanisms of action to support its potential use in managing acute mountain sickness (AMS). We first established and validated an HPLC method for quality control, then comprehensively profiled the chemical composition using LC-MS. Network pharmacology and molecular docking were applied to predict the core anti-hypoxic components, candidate targets and signaling pathways. The primary bioactivity was further verified through an in vitro carbonic anhydrase (CA) inhibition assay. A total of 71 constituents were identified, with kaempferol and ellagic acid emerging as the primary anti-hypoxic phytochemicals. These compounds target seven core proteins (SRC, PIK3R1, ESR1, EGFR, PTK2, IGF1R, and LYN) to regulate vascular tone, inflammation, oxidative stress, blood–brain barrier integrity, and cell survival under hypoxic conditions. By modulating pathways such as HIF-1α, PI3K/AKT, FAK/PTK2, SRC, and IGF1R, these phytochemicals ultimately influence the onset and alleviation of AMS. Enzyme inhibition assays demonstrated that kaempferol and ellagic acid inhibited CA with IC50 values of 34.05 μM and 119.1 μM, respectively. Molecular docking further revealed that both compounds suppressed CA activity through a combination of hydrogen bonding and hydrophobic interactions, consistent with a zinc-bound water-anchoring mechanism. This study elucidates the phytochemical basis and molecular mechanism responsible for the anti-hypoxic effects of GSHJT Oral Liquid, providing scientific support for its potential application as a natural, plant-derived intervention for preventing and alleviating acute mountain sickness, providing scientific support for its potential application and offering a reproducible paradigm for the rational development of other Rhodiola-based phytomedicines, though further in vivo validation is required to confirm the anti-hypoxic efficacy.