The objective of this work is to explore the phytochemical profile of Mentha piperita essential oils (MPEO) collected from two different Moroccan regions using gas chromatography-mass spectrophotometer (GC-MS) and to investigate their antioxidant, anti-inflammatory, antidiabetic and, antimicrobial effects using in vivo and in vitro assays. The chemical constituent of MPEO from the Azrou zone is dominated by carvone (70.25%), while MPEO from the Ouazzane zone is rich in Menthol (43.32%) and Menthone (29.4%). MPEO from Ouezzane showed higher antioxidant activity than EO from Azrou. Nevertheless, EO from Ouezzane considerably inhibited 5-Lipoxygenase (IC50 = 11.64 ± 0.02 µg/mL) compared to EO from Azro (IC50 = 23.84 ± 0.03 µg/mL). Both EOs from Azrou and Ouazzane inhibited the α-amylase activity in vitro, with IC50 values of 131.62 ± 0.01 µg/mL and 91.64 ± 0.03 µg/mL, respectively. The EOs were also tested for minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). The discdiffusion test revealed that MPEOs from both regions have significant antibacterial efficacy, and MPEOs from the north region showed the highest effect. The gram-positive bacteria were the most susceptible organisms. The MIC concentrations were in the range of 0.05 to 6.25 mg/mL, and the MBC concentrations were within 0.05–25.0 mg/mL. The MBC/MIC index indicated that MPEO has strong bactericidal effects.