With the growing issues of food spoilage, microbial resistance, and high mortality caused by cancer, the aim of this study was to evaluate T. zygis essential oil (TZEO) as a potential solution for these challenges. Here, we first performed GC/MS analysis which showed that the tested TZEO belongs to the linalool chemotype since the abundance of linalool was found to be 38.0%. Antioxidant activity assays showed the superiority of TZEO in neutralizing the ABTS radical cation compared to the DPPH radical. The TZEO was able to neutralize 50% of ABTS•+ at the concentration of 53.03 ± 1.34 μg/mL. Antimicrobial assessment performed by employing disc diffusion and minimal inhibitory concentration assays revealed TZEO as a potent antimicrobial agent with the highest inhibition activity towards tested gram-negative strains. The most sensitive on the treatment with TZEO was Enterobacter aerogenes showing an MIC 50 value of 0.147 ± 0.006 mg/mL and a MIC 90 value of 0.158 ± 0.024 mg/mL. Additionally, an in situ analysis showed great effects of TZEO in inhibiting gram-negative E. coli, P. putida, and E. aerogenes growing on bananas and cucumbers. Treatment with the TZEO vapor phase in the concentration of 500 μg/mL was able to reduce the growth of these bacteria on the food models to the extent > 90%, except for E. coli growth on the cucumber, which was reduced to the extent of 83.87 ± 4.76%. Furthermore, a test on the antibiofilm activity of the tested essential oil revealed its biofilm prevention effects against Salmonella enterica which forms biofilms on plastic and stainless-steel surfaces. Performed tests on the TZEO effects towards cell viability showed no effects on the normal MRC-5 cell line. However, the results of MTT assay of TZEO effects on three cancer cell lines (MDA-MB-231, HCT-116, and K562) suggest that TZEO exerted the strongest effects on the inhibition of the viability of MDA-MB-231 cells, especially after long-term treatment in the highest concentration applied with reducing the viability of the cells to 57%. Additionally, results of NBT and Griess assays suggest that TZEO could be a convenient candidate for future testing for developing novel antitumor therapies.