Probiotics, according to the World Health Organization's (WHO) definition, refer to live microorganisms that confer health advantages when ingested in appropriate quantities. These benefits encompass enhancements in gut health, immune function, infection prevention, and cholesterol management. This investigation shifts its focus towards kombucha, a fermented tea beverage gaining prominence for its conceivable health merits. Its assorted microbial composition, encompassing acetic acid bacteria, lactic acid bacteria, and yeasts, is instrumental in bestowing it with probiotic attributes. The study's primary goals involve the creation of kombucha concoctions using varied nutritional sources such as Chia seeds, intact and ground sativa seeds, and black tea. The aim was to isolate lactic acid bacteria with potential probiotic qualities. This pursuit resulted in the successful isolation of 29 bacterial strains based on acid production. Subsequent morphological and fundamental physiological assessments of these 29 strains led to the identification of only 4 strains that fulfilled the basic probiotic criteria, while those exhibiting blood hemolysis and producing catalase, oxidase, and DNase were excluded. The four isolates coded as E3 from black tea and C9 (1-3) from ground sativa seed. Among these 4 strains, their potential to withstand gastric and intestinal conditions was examined. The strain labeled as E3 exhibited moderate tolerance to gastric conditions, particularly at pH 3, but showed intolerance to intestinal conditions. Conversely, the remaining strains displayed no tolerance towards either gastric or intestinal conditions. However, all isolates demonstrated tolerance to varying concentrations of bile (0.3-2%). Upon identification, it was established that the genus of the four strains was Lactobacillus. Specifically, the E3 isolate was identified as Lactobacillus plantarum with a 99.9% identification match. The isolates C9 (1-3) were categorized as Lactobacillus paracasei in three strains (43.6%, 45.2%, and 44.9% identification match, respectively). According agar well diffusion method the antibacterial potential of the filtrate produced by the 4 isolates exhibited differing levels of inhibition against the target bacteria. E3 demonstrated the most robust activity, primarily against Bacillus cereus, with notable inhibition against Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, and Lactobacillus plantarum. Remarkably, isolate C9 (3) displayed broad-spectrum antibacterial efficacy against all tested strains except Escherichia coli, where no activity was observed. Conversely, the filtrate of C9 (2) displayed no antimicrobial efficacy, while the filtrate of C9 (1) demonstrated heightened activity against Pseudomonas aeruginosa compared to Bacillus cereus and Staphylococcus aureus. The spot method yielded analogous results, except that C9 (2) exhibited activity against Bacillus cereus and Escherichia coli. Furthermore, when the disc diffusion method was employed, the four isolates demonstrated considerable susceptibility to various antibiotics. The antibiotic susceptibility profiles offered insights into the isolates' responsiveness to distinct antibiotics. In conclusion, while the anticipated probiotic potential wasn't entirely realized, this study paves the way for the discovery of antimicrobial compounds with potential against multidrugresistant bacteria. It enriches our comprehension of probiotics, particularly in the context of kombucha, and foreshadows future investigations into their health-promoting capacities.