In recent years, there has been great development in the search for new natural compounds for food preservation aimed at a partial or total replacement of currently popular antimicrobial chemicals. Carob (Ceratonia siliqua) offers a natural promising alternative for food safety and bioconservation. In this work, the methanolic extract of carob leaves (MECL) was tested for the ability to inhibit the growth of a range of microorganisms. MECL inhibited the growth of Listeria monocytogenes at 28.12 μg/mL by the broth microdilution method. The effect of this bacteriostatic concentration on the growth of this bacterium revealed a pattern of inhibition characterized by (a) a resumed growth phase, which showed a lower rate of growth if compared with controls; and (b) first a lag and then a stationary phase at a lower bacterium concentration. The study of the chemical composition of MECL by high-performance liquid chromatography and liquid chromatography/mass spectrometry showed the presence of gallic acid, (-)-epigallocatechin-3-gallate, myricitrin, isoquercitin, catechin, chlorogenic acid, and malic acid. L. monocytogenes growth inhibition was recorded for myricitrin and gallic acid at 450 μg/mL and for (-)-epigallocatechin-3-gallate and isoquercitin, respectively, at 225 and 112.5 μg/mL. Taking into account that proline is a ligand of proline dehydrogenase (PDH), the use of this compound leads us to hypothesize the mode of action of MECL constituents.