Waste lignocellulosic biomass obtained from the dry plant matter is the most abundantly available resource for the production of biofuels, and biochar. The invasive weed tree of Prosopis juliflora was employed as feedstock for the extraction process, which converts biomass into biogas, bio-oil, and biochar in the presence of subcritical water at high temperatures (250 °C to 374 °C) and pressures (4-22 MPa). The extraction process was performed inside a 50 ml stainless steel hydrothermal reactor with 3.5 g of feedstock and varying process parameters such as temperatures (250–325 °C) and reaction time (30–120 min) and biomass to water loading (10–30 % w/v). The response surface methodology was employed to optimize the parameters for maximizing the bio-oil yield under subcritical condition using Design Expert 8.0.7.1 software. The % yield of bio-oil and biochar during this process were taken as responses. The biomass and bio-oil were characterized using proximate and ultimate elemental analysis, thermogravimetric analysis, and gas-chromatography mass spectroscopy. The results showed that the maximal yield of bio-oil 3.65 % was obtained at a temperature of 277.62 °C, reaction time 59.98 min and biomass to water loading 20.13 % w/v. The resulted bio-oil was found to contain long-chain alkanes, ketones, carboxylic acids, amines, and phenols.