In this paper, hydrothermal liquefaction (HTL) of microalgae Chlorella pyrenoidosa was conducted for biocrude oil production. The ethanol water co-solvent was introduced to take advantage of the special properties of supercritical ethanol and hot compressed water. The effects of the ethanol/water mass ratio (R) on the product distribution and characterization were discussed. The co-solvent showed better performance over any constituent mono-solvent, indicating synergistic effects of ethanol and water. The highest biocrude oil yield of 57.3% and the lowest solid residue yield of 9.4% were achieved at R = 5:2. The loading factor was considered as another influencing factor under supercritical conditions. Furthermore, potential reaction pathways for major biomacromolecule monomers in the ethanol water co-solvent were also proposed on the basis of the gas chromatography mass spectrometry (GC-MS) analysis of the liquid products.