In Vitro Study of the Antimicrobial and Prebiotic Activity of Origanum heracleoticum Hydrolates from Two Italian Crops and the Proteomic Response of Probiotic Bacteria

This study explores the chemical composition and biological activities of essential oils (EOs) and hydrolates (Hys) from Origanum heracleoticum (OH), an aromatic plant with known applications in the food industry. While EOs have been widely studied, Hys (co-products of EO distillation) remain largely overlooked. OH was cultivated in the Lucanian and Casola areas, respectively in the regions of Basilicata and Emilia-Romagna, to evaluate the influence of geographic and climatic factors on product composition and activity. Gas chromatographic analysis showed significant variability between the EO and its related Hy. Carvacrol emerged as the dominant compound, ranging from 71% to 91% in both extracts followed by γ-terpinene and thymol in EOs and Hys, respectively. Both EOs and Hys exhibited antimicrobial activity against tested bacterial strains even if active compounds of Hys were relatively more effective than that of EOs. Notably, Lactobacillus rhamnosus (probiotic) showed greater resistance to EOs compared to Escherichia coli (pathogenic), whereas Hys inhibited L. rhamnosus at 25% v/v, while no MIC against E. coli was reached within the tested concentration range (>50% v/v). Interestingly, sub-inhibitory concentrations of Hys (especially those from Lucan area) stimulated L. rhamnosus growth, promoting earlier exponential phase entry and higher final cell densities. Proteomic analysis revealed associated changes in protein secretion, suggesting adaptive metabolic responses. These findings highlight the distinct chemical and biological properties of Hys obtained from a spice widely used in food, and their potential as functional agents. Their ability to influence L. rhamnosus growth in vitro supports further investigation of their potential microbiota-modulating properties and the development of food supplements and microbiome-friendly formulations.