An investigation of nanoemulsions of tea tree (Melaleuca alternifolia) oil and its main constituent terpinen-4-ol was carried out compared to non-formulated products for their antimicrobial and antioxidant properties. To prepare nanoemulsions from coarse oil-in-water (O/W) emulsions, the products were added to aqueous solutions containing 10% Tween 80 and stirred continuously under ultrasonication. Interestingly, the tea tree oil nanoemulsion produced droplets with a size of 70.67 nm, while the terpinen-4-ol nanoemulsion produced droplets with a size of 176.00 nm. The nanoemulsions showed high negative zeta potentials (-22.30 and -23.50 mV for oil and terpinen-4-ol, respectively. Compared with tea tree oil and terpinen-4-ol nanoemulsions as antioxidants, terpinen-4-ol nanoemulsion had the highest radical scavenging activity against DPPH with the lowest EC50 value (253.65 mg/L). Furthermore, terpinen-4-ol nanoemulsion exhibited the highest antibacterial activity (MIC = 600 and 475 mg/L against Salmonella typhimurium and Staphylococcus aureus, respectively). Tee tree oil nanoemulsion, on the other hand, had the strongest inhibition effect against Candida albicans (EC50 = 74.86 mg/L). Furthermore, a nanoemulsion of terpinen-4-ol exhibited high antifungal activity (EC50 = 253.13 and 312.53 mg/L against Aspergillus flavus and A. niger, respectively). Despite this, tea tree oil nanoemulsion was the most effective against fungal spores (EC50 = 484.61 and 291.99 mg/L against spores of A. flavus and A. niger, respectively). The information in this study will lead to a potential breakthrough for eco-friendly antimicrobials and antioxidants that contain tea tree oil and terpinen-4-ol.