Optimization of three-component essential oil-loaded nanoemulsions in combination using D-optimal mixture design

Abstract

This study aimed to select an optimal combination of Rosmarinus officinalis, Salvia officinalis, and Thymus satureioides nanoemulsions to overcome the resistance of some pathogenic bacterial strains and inhibit oxidative stress. For this purpose, the chemical composition of essential oils (EO) obtained from these herbs was determined using gas chromatography/mass spectrometry (GC/MS). Oil-in-water (O/W) nanoemulsions were prepared and characterized. The antioxidant activity of the nanoemulsions and their combinations was evaluated, while the antibacterial activity was determined against two Gram-negative and two Gram-positive bacteria. The optimal mixture of nanoemulsions was determined using Design Expert software. GC/MS analysis revealed that R. officinalis EO was dominated by 1,8-cineole (39.48%). Endo-borneol (20.73%) and thymol (15.16%) were found as the main compounds of T. satureioides EO, while S. officinalis EO was mainly dominated by caryophyllene (30.49%). The prepared nanoemulsions' mean droplet sizes oscillated between 30.99 and 321.30 nm, whereas PdI was found to be 0.549 for R. officinalis, 0.809 for S. officinalis, and 0.209 for T. satureioides. For antioxidant activity, the combination composed of 34.11% rosemary nanoemulsion, 24.11% sage nanoemulsion, and 41.77% thyme nanoemulsion revealed the highest capacity. Regarding antibacterial activity, the optimal mixture consisting of 41.90% R. officinalis nanoemulsion, 17.30% S. officinalis nanoemulsion, and 40.90% T. satureioides nanoemulsion showed the strongest antibacterial effect against E. coli, S. aureus, and E. faecalis, while the optimal effect against P. aeruginosa was predicted using a combination containing 43% sage nanoemulsion and 57% thyme nanoemulsion.