Thermochemical catalytic liquefaction (TCL) of the marine microalgae Dunaliella tertiolecta was performed in ethylene glycol acidified with H2SO4 as a catalyst. The mathematical model for predicting the liquefaction yield was set up by a central composite rotatable design (CCRD) and response surface analysis (RSA). A total of 23 individual experiments were conducted to study the effect of H2SO4 concentration, reaction temperature, and reaction time on the liquidation yield. From a regression analysis. the conversion yield of microalgae cells into liquid is simply expressed as a function of the operating variables by polynomial containing quadratic terms. The highest liquefaction yield of microalgae would be 97.05%, at the following optimized reaction conditions: an amount of added H2SO4 of 2.4%, and a reaction temperature of 170 degrees C, with a 33 min reaction time. To put bio-oils into wide application. the various physical and chemical characteristics of bio-oils at the conditions of the maximum product yields were determined, and the detailed chemical compositional analysis of bio-oils was performed by various spectroscopic techniques Such as Fourier transform infrared spectroscopic analysis (FT-IR), carbon-13 nuclear magnetic resonance (C-13 NMR), and gas chromatography - mass spectrometry (GC-MS). The bio-oils were composed of benzofuranone, fatty acid methyl ester, and fatty acid hydroxyethyl ester, with a long chain from C-14 to C-18. These bio-oils were presented its an environmentally friendly feedstock candidate for biofuels and chemicals.