Development of microemulsion delivery systems for bioactive compounds : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Food Technology at Massey University, Auckland, New Zealand

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Many bioactive compounds for health benefits are not readily stable against degradation and their solubility is also very low. As a result, a delivery system is required to encapsulate and protect bioactive compounds for their food applications. Emulsion is one of the delivery systems which has been studied by many researchers. But emulsion tends to destabilize during storage and its opaque optical properties makes it difficult for its use and incorporation into clear foods or beverages without affecting their original appearance. Therefore, microemulsion, which is known to be transparent, has been investigated to some extent to encapsulate and deliver bioactive compounds as a potential delivery system. The objective of this research was to fabricate oil-in-water (O/W) microemulsions which might be utilised as the delivery system for bioactive compounds. This thesis is mainly composed of two sections. The first section was to produce microemulsions via emulsion dilution method and water titration method as well as to study the characteristics of these microemulsions. Beta-carotene was a type of bioactive compound used in the second section to study the effect of beta-carotene on the formation and properties of microemulsion which was fabricated using the same methods described above. At first, emulsion dilution method was employed to fabricate microemulsions with different types and concentrations of oils, such as peanut oil, fractionated coconut oil, isopropyl myristate (IPM), lemon oil and Capmul 708G, and also with different surfactants (Tween 20, 40, 60 and 80). It was found that peanut oil and fractionated coconut oil could not be utilised to form microemulsions by this method, whereas IPM and lemon oil had the ability to fabricate microemulsions. When 1% Tween 80 was introduced as the surfactant and dilution medium, microemulsion could be formed when the concentration of IPM was less than 0.1% and that of lemon oil was less than 0.2%. Among the different types of Tween surfactants, Tween 80 was the most efficient when its solution containing Tween micelles was used as a dilution medium compared to the other Tween surfactants because more lemon oil could be incorporated into the Tween 80 micelles with an increase in Tween 80 concentration. In the following study, a water titration method was employed to create ternary or pseudo phase diagrams which indicated the ability to fabricate microemulsions of a mixture system. Various types of oils (Captex 100, Capmul PG-8, Capmul PG-12, Capmul PG-2L, lemon oil, Capmul MCM C8, Capmul 708G and Captex 355) and surfactants (Tween 80, Tween 20, Span 80 and Kolliphor EL) were used in this study. Absolute ethanol and propylene glycol (PG) were also incorporated as cosurfactant and cosolvent, respectively. It is concluded that all these oils and surfactants could be utilised by the water titration method to produce microemulsions, however, their ability to form microemulsions were different. Capmul 708G, which is a monoglyceride, was the most efficient in terms of producing microemulsions compared to diglyceride and triglyceride. Tween 20 and Kolliphor had the similar emulsifying properties compared to Tween 80 whereas Span 80 was not efficient. Both absolute ethanol and PG could assist the formation of microemulsions when they were introduced into the mixture system of oil, surfactant and water. In the following study, microemulsions containing 0.1% and 0.4% lemon oil and an emulsion containing 1.5% lemon oil (larger oil droplets), which were fabricated by the emulsion dilution method, were chosen to incorporate beta-carotene as a lipophilic model bioactive compound into lemon oil in order to study its impact on the formation and properties of the resulting microemulsion and emulsion systems. The encapsulation of beta-carotene into 0.1% and 0.4% lemon oil caused a significant increase in the particle size of the O/W microemulsions, but the particle size was still within the size range of microemulsion. As a result, the beta-carotene-loaded microemulsions containing 0.1 and 0.4% lemon oil were visually clear in appearance. However, the incorporation of beta-carotene did not increase and alter the particle size of the emulsion containing 1.5% lemon oil. The microemulsion sample containing 0.1% lemon oil and the emulsion containing 1.5% lemon oil were stored at 25 °C without exposed to oxygen and light for one month. While, the microemulsion containing 0.4% lemon oil was selected and placed at three different temperatures (4, 25 and 37 °C) for 1 month: at 4 and 37 °C without exposure to both oxygen and light and at 25 °C, four different environmental conditions (i.e. with oxygen/light, with oxygen and without light, without oxygen and with light, without oxygen/light). The results showed that the rate of beta-carotene degradation was lower in all these three samples when compared to the beta-carotene present in a hexane solution without encapsulation. Higher temperature accelerated the degradation rate of beta-carotene. As a consequence, the 0.4% lemon oil microemulsion at 4 °C exhibited the slowest degradation rate of beta-carotene. Next, the microemulsions fabricated by the water titration method were selected to encapsulate beta-carotene to study the encapsulation capacity of these microemulsion systems as well as their ability to protect beta-carotene against oxidative degradation during storage. Capmul 708G, Tween 80, Milli-Q water and PG mixture system were chosen to fabricate microemulsions and two formulations (L910 and L990) were prepared to incorporate beta-carotene. L910 was comprised of 81% Capmul 708G, 9% Tween 80, 5% water and 5% PG, whereas L990 contained 9% Capmul 708G, 1% Tween 80, 45% water and 45% PG. It was able to see clearly from this experiment that the L910 system could incorporate more beta-carotene than L990. Both L910 and L990 could reduce the degradation rate of beta-carotene when loaded into them compared to their presence in hexane solutions without encapsulation. Similar to the previous experiment as described above, when the beta-carotene incorporated microemulsions were placed at 4 °C and away from oxygen and light, beta-carotene had the highest retention rate after storage for 1 month. Furthermore, beta-carotene degradation rate in L910 was slower than that in L990, indicating L910 was more effective than L990 in terms of incorporating and protecting beta-carotene. It is shown clearly from the present study that microemulsions could be formed via the water titration and emulsion dilution methods. The type and concentration of oil phase and surfactant had a significant influence on the determination of whether a mixture system could form a microemulsion as well as the properties of the formed microemulsion. The microemulsions produced by these two different methods could be utilised to encapsulate beta-carotene as the incorporation of beta-carotene did not have a significant influence on the properties of the original microemulsions. Moreover, microemulsions provided the stability and protection to beta-carotene against oxidative degradation that could be caused by oxygen, light and temperature during storage, which might be possible to be applied to some liquid foods and beverages.
Bioactive compounds, Microencapsulation, Emulsions, Functional foods