Formation And Characterization Of Antimicrobial And Antioxidant Thymol Microemulsion Systems

Thymol (5-methyl-2-isopropyl phenol) is the major component of essential oil prepared from the herb thyme (Thymus vulgaris). The antimicrobial and antioxidant effectiveness of thymol have been known for decades. However, the application of thymol in food products is limited for its low water solubility. Micremulsification is a good way to disperse hydrophobic molecules in aqueous phase.Pseudo ternary phase diagrams were constructed to assess the ability for microemulsion formation and dilutability of each system using non-ionic, cationic, and anionic surfactants. The influence of the surfactants and ethanol on the phase behavior of the microemulsion system was reflected in pseudo-ternary diagrams. The microstructure transition of a U-type microemulsion along the dilution line at the 90/10 surfactant/oil mass ratio, Tween 80 (surfactant) and thymol and ethanol (3:1, w/w) (oil phase) was explored. Differential scanning calorimetry (DSC) reveals the states of water at different aqueous phase content and suggests that dispersion phases gradually inverted from the w/o (0-20% water) to bicontinuous phase (25-35% water), and finally o/w phase (40-90% water) along the dilution line. The transition points detected by conductivity measurments coincide with the result of DSC measurements. Whereas, the maximum point of viscosity shows not at bicontinuous region, but at 60% aqueous phase system, which suggests the formation of hexagonal liquid crystalline. On the basis of results of DSC, conductivity, and viscosity, we proposed a possible structural change of the U-type microemulsion along the dilution, which are inverse micellar phase, amorphous phase, bicontinuous phase, hexagonal liquid crystalline, and normal micellar with increasing aqueous phase content.The antimicrobial activity assay using kinetics of killing analysis demonstrated that the microemulsions in w/o regions exhibited relatively high antimicrobial activity against Escherichia coli and Staphylococcus aureus due to the antimicrobial oil phase as the continuous phase, while the antimicrobial activity started to decrease when the microemulsions entered the bicontinuous region, and decreased rapidly as the water content increased in the o/w region, as a result of the dilution of antimicrobial oil droplets in the aqueous continuous phase. DPPH scavenging activity of thymol showed an increasing trend with the increase of water content. Although higher content of thymol in W/O microemulsion, a lower DPPH scavenging activity was observed due to the hinderance of Tween 80. Formation of larger surface area in O/W microemulsion may contribute to the good DPPH scavenging activity of thymol.Surfactants are generally used in food systems at low concentration, therefore we studied the critical thymol concentration (Csat) of 2.0%(w/w) Tween 80 solution and the Csat of 0.2% was get.’H NMR spectra indicated that thymol accommodate near the ester group between the polyoxyethylene headgroup and the oleic acid in Tween 80 micelle. The antioxidant activity of thymol in ethanol and thymol micelle was evaluated by DPPH scavenging activity and Ferric reducing antioxidant power, and compared with synthetic antioxidant (BHT). Thymol in micelle showed slightly higher antioxidant activity than thymol in ethanol. Both of the method exhibited similar antioxidant order:BHT> thymol in micelles> thymol in ethonal. The antioxidant activity of thymol micelle is comparable to that of BHT, which suggest that thymol is a potential natural antioxidant in food system.