Tobacco cultivation is characterized by high amounts of waste biomasses whose disposal frequently represents a complex and expensive problem. A study was conducted to evaluate thepotential of pre-harvest light air-cured (Burley) and dark fire-cured (Kentucky) tobacco waste biomasses as a source of bioactive compounds (nutraceutical ingredients) such as polyphenols. Pre-harvest waste materials (topping fresh materials and residual stalks at final harvest) were collected to determine dry matter, total polyphenols content (TPC; Folin assay), and DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,20-azino-bis(3-ethylbenzothiazoline-60-sulfonic acid) diammonium salt) antioxidant capacity. Polyphenols quali-quantitative profiles obtained by Orbitrap Q Exactive of both tobacco types were also determined. Total pre-harvest waste biomass amounted to 3956.9 and 1304.4 kg d.w. ha−1 in light air-cured (Burley) and dark fire-cured (Kentucky) tobacco types, respectively. Polyphenols content, expressed as g kg−1 dry weight (d.w.), ranged between 4.6 and 15.7 g kg−1 d.w. and was generally greater in leaves than in stalks. Considering both leaves and stalks, the light air-cured (Burley) tobacco crop yielded 22.1 kg ha−1 of polyphenols, while the dark fire-cured (Kentucky) tobacco yielded 12.0 kg ha−1. DPPH and ABTS were significantly greater in leaves than in stalks waste biomass in both types of tobacco. The most abundant components were quinic and chlorogenic acids, rutin, and luteolin rutinoside.