Abstract: The emergence of antibiotic resistant bacteria has made the treatments of pathogenic infections difficult. The discovery that some analytes of plant extracts are active against multi-drug resistant bacteria has opened ways to researching into the antimicrobial activity of these phytocomponents. Studies are designed to determine the antimicrobial and antioxidant potentials of the volatile oil of Ocimum gratissimum and its inhibition on partially purified and characterized extracellular protease of Salmonella enteritidis. The oil was extracted by hydrodistillation. Its antimicrobial effect was carried out using microdilution method while antioxidant effect was tested against 1,1-diphenyl-2-picrylhydrazyl (DPPH). The caseinolytic activity of the enzyme was studied with oil as inhibitor. The enzyme was partially purified with dialysis and gel filtration. The oil has scavenging activity of 66.98±1.78% compared to butylated hydroxyltoluene (positive control), 53.29±2.51% (sig. p<0.0001) against DPPH. Salmonella enteritidis was inhibited by this oil with IC50 of 3.98% (v/v). The enzyme had optimal activities at 45°C and pH 7.5. None of the metallic chloride tested produced any significant increase in the activity of the enzyme. Hg2+ and Pb2+ were inhibitors of this enzyme. The oil showed a non-competitive inhibition with Km = 0.33 mg mL-1, Vmax = 1.25x103 μmol min-1 (oil absent) and V’max = 2.50x102 μmol min-1 (oil present). The highest purification fold = 1.88 and the highest percentage yield = 51.74 as compared to the crude enzyme extract. Therefore, the volatile oil of Ocimum gratissimum possessed antimicrobial and antioxidant activities, its inhibition against this protease may be one of the ways of its antimicrobial effect.