The Mentha genus is a well-known medicinal plant and is found to contain various phenolic compounds. Most of these compounds are typically extracted; however, utilizing biotechnology methods, such as hairy root culture, offers a viable option for producing these compounds. Hairy roots were generated from four Mentha species, including Mentha spicata, Mentha piperita, Mentha aquatica, and Mentha longifolia. Afterward, the phenolic content and antioxidant properties of both parts were compared. The results indicated that both the type of explant and the species studied are crucial factors in determining the types and levels of phenolic compounds present. It was also found that the highest total phenol and flavonoid contents were related to the aerial parts extracts of M. aquatica and M. spicata, while these contents were higher in the hairy roots of M. piperita and M. longifolia. Moreover, the HPLC analysis of the aerial parts extracts of different Mentha species showed that the highest levels of caffeic acid, chlorogenic acid, rosmarinic acid, and quercetin were found in M. spicata. However, in the case of hairy roots, M. piperita extract showed the highest amounts of these compounds. In general, the amount of secondary metabolites in the hairy root extract was lower than in the aerial parts. Additionally, the highest level of antioxidant properties was found in the leaf extracts. In conclusion, the aerial parts of the Mentha species have more phenolic compounds than the hairy roots under conditions where no inducers or growth stimulants are used.