Mentha gattefossei Maire (Lamiaceae) is a perennial and aromatic plant, endemic to Morocco [1]. Its aerial parts are used traditionally to alleviate symptoms of the digestive disorders, bronchitis, cold, cough and migraine. Also, the plant is used as food and for flavoring purposes [1, 2]. This study assessed the polyphenolic profile and antioxidant activity of an alcoholic extract obtained from aerial parts of M. gattefossei species cultivated in Republic of Moldova within ex situ conservation programmes at international level. The extract of M. gattefossei was subjected to HPLC-DAD-ESI-Q-TOF-MS/MS analysis and separated on a Phenomenex Gemini C18 (100 × 2 mm, 3 μm) column, with a gradient obtained from 0.1% formic in water and 0.1% formic acid in acetonitrile. The levels of total phenolics and flavonoids were also estimated. The extract showed potent reducing capacity and free radical scavenging properties and also good iron-chelating activity. The metabolite profiling revealed the presence of several phenolic acids (hydroxybenzoic acid, chlorogenic acid, rosmarinic acid, salvianic acid A, syringic acid) and flavonoids (diosmin, isoquercitrin, acacetin rutinoside). Of these, rosmarinic acid was by far the major phenolic compound, its content reaching 12.42 ± 0.23 mg/g extract. This investigation provided an insight on polyphenols from Mentha gattefossei and their bioactivity.