Mycobacterium tuberculosis is one of the most common causes of morbidity and mortality among adults who have tested HIV-positive or those living in poverty. One of the major reasons for the need of extended chemotherapeutic regimens and widespread epidemicity of tuberculosis is that the causative agent, M. tuberculosis, has the ability to be in a dormant state. Therefore, there is an urgent need for new lead compounds that are effective against M. tuberculosis in both its active and dormant states. In this study, we investigated the anti-mycobacterial activity of the total ethanol extract and fractions of Melissa officinalis L. (Lemon balm) under both actively growing and dormancy inducing conditions. As a result, the total extract of M. officinalis exhibited antimicrobial activity against both active and dormant states of Mycobacterium smegmatis and Mycobacterium bovis BCG with Minimum Inhibitory Concentration (MIC) values ranging from 12.5 µg/mL to 50 µg/mL. Among the fractions, the n-hexane fraction showed the most ideal activity against active and dormant states of M. smegmatis (MIC = 6.25 µg/mL). In addition, the n-hexane fraction also exhibited antimicrobial activity against actively growing M. bovis BCG (MIC = 12.5 µg/mL), while moderate activity (MIC = 50 µg/mL) was observed against the dormant state of M. bovis BCG.