Chlorpromazine (CPZ) was exposed to a 266 nm laser beam for different periods of time ranging from minutes to 24 h. At intervals, the products from irradiation were evaluated by thin-layer chromatography (TLC) and evaluated for their activity against mycobacteria of human interest (Mycobacterium tuberculosis, M. avium, M. intracellulare and their corresponding reference strains or clinical isolates). With the exception of the M. avium 47/07 clinical strain, the products produced from the irradiation of CPZ for 4 h had greater activity against M. intracellulare ATCC, M. avium ATCC, H37Rv and the Multidrug-resistant tuberculosis (MDR-TB) strains as opposed to that produced by the unirradiated control. The level of products from the 4-h exposure of CPZ remained the same throughout the next 20 h of irradiation. Of significant note is that the irradiation products of CPZ had lower in vitro cytotoxicity against human cells, suggesting that this approach may be useful for the development of compounds more bioactive than the parental species.