Hydrophobic deep eutectic solvents (HDESs) were first employed under hydrothermal conditions to form a two-phase system for instantaneous and efficient lipid extraction from wet microalgae, preventing lipid degradation and avoiding the use of toxic solvents. Ten HDESs were assessed for their efficiency and selectivity in lipid extraction. Among them, OCT-DEC, composed of octanoic acid and decanoic acid, achieved the highest extraction efficiency at 95.42 %, while also preventing the co-extraction of proteins and reducing sugars. Molecular dynamics simulations revealed that lipid extraction efficiency is primarily influenced by lipid-HDES interactions, which drive lipid diffusion, and the interfacial properties of the two-phase hydrothermal system, which resist lipid diffusion. The superior performance of OCT-DEC is attributed to its high hydrogen bond density and short bond lifetimes, which, combined with its structural stability, synergistically reduce resistance to lipid diffusion at the interface. Additionally, OCT-DEC significantly enhances the thermal stability of unsaturated fatty acids, such as C20:5, effectively preventing the degradation of high-value lipids and highlighting its potential for hydrothermal extraction of these valuable compounds.