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The essential oil of <i>Jasminum grandiflorum</i> has very good biological activity. The present study aimed to analyze the chemical composition and biological activity of <i>J. grandiflorum</i>. The main constituents of <i>J. grandiflorum</i> essential oil were benzyl acetate (37%), benzyl benzoate (34.7%) and linalool (9.6%). The antioxidant activity was 58.47%, which corresponds to 220.93 TEAC. The antimicrobial activity was weak to moderate, with inhibition zones ranging from 2.33 to 5.33 mm. The lowest MIC value was against <i>Candida glabrata</i>. The antimicrobial activity of the vapor phase of the essential oil was significantly stronger than that of the contact application. Biofilm analysis using a MALDI-TOF MS Biotyper showed changes in the protein profile of <i>Pseudomonas fluorescens</i> that confirmed the inhibitory effect of <i>J. grandiflorum</i>. The insecticidal potential of the essential oil against <i>Oxycarenus lavatera</i> and <i>Brassicogethes aeneus</i> was also demonstrated. Due to the properties of <i>J. grandiflorum</i> essential oil, it could find application as a biofilm control agent for the shelf-life extension and storage of fruits and vegetables and as a possible insecticidal agent.