Article pubs.acs.org/JAFC
Increased Antioxidant Efficacy of Tocopherols by Surfactant Solubilization in Oil-in-Water Emulsions S. Sezer Kiralan,*,†,‡ Esra Doğu-Baykut,†,§ Ketinun Kittipongpittaya,∥ David Julian McClements,†,⊥ and Eric A. Decker†,⊥ †
Department of Food Science, University of Massachusetts Amherst, 240 Chenoweth Laboratory, 100 Holdsworth Way, Amherst, Massachusetts 01003, United States ‡ Department of Food Engineering, Faculty of Engineering, Ankara University, 06110 Ankara, Turkey § Department of Food Engineering, Faculty of Chemical and Metallurgical Engineering, Istanbul Technical University, Ayazaga Campus, Maslak, 34469 Istanbul, Turkey ∥ Department of Agro-Industry Technology and Management, Faculty of Agro-Industry, King Mongkut’s University of Technology North Bangkok, Bangkok 10800, Thailand ⊥ Bioactive Natural Products Research Group, Department of Biochemistry, Faculty of Science, King Abdulaziz University, Post Office Box 80203, Jeddah 21589, Saudi Arabia ABSTRACT: The physical location of antioxidants in oil-in-water emulsions can have significant influence on their free radical scavenging activity and ability to inhibit lipid oxidation. We aimed to determine the effect of the surfactant concentration on the partitioning behavior of tocopherols (α, γ, and δ) in oil-in-water emulsions. Tween 20 (0.1, 0.5, and 1%) increased the partitioning of the tocopherols into the aqueous phase via the formation of Tween 20−tocopherol comicelles. Partitioning behavior of antioxidants was dependent upon the number of methyl groups and, thus, polarity of the tocopherols. δ-Tocopherol (one methyl group) exhibited the most partitioning into the aqueous phase, while α-tocopherol (three methyl groups) had the lowest partitioning. Lipid oxidation studies showed that the antioxidant activity of δ- and α-tocopherols was enhanced by adding Tween 20 to oil-in-water emulsions. This work suggests that surfactant micelles could increase the antioxidant activity of tocopherols by changing their physical location. KEYWORDS: antioxidant, oil-in-water emulsion, surfactant, tocopherol, lipid oxidation
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INTRODUCTION In food products, lipids are susceptible to oxidative deterioration, leading to the development of undesirable color, texture, and rancidification. Food emulsions are one of many foods that are affected by lipid oxidation reactions.1 The oxidation of lipids causes food quality deterioration and reduces the nutritional value of foods. In addition, toxic reaction products are formed, and they can induce health problems, such as cancer and neurodegenerative diseases.2 The addition of natural or synthetic antioxidants in foods can control the rate of lipid oxidation by scavenging free radicals or chelating metals.1 Many factors, such as chemical reactivity, environment in which the antioxidant partitions, and physical properties of the food, may affect the activity of antioxidants in heterogeneous food systems, such as oil-in-water (O/W) emulsions. In many foods, the most effective factor impacting the activity of free radical scavengers is the physical location of the antioxidant, which is dependent upon their polarity.3 Porter and co-workers4 evaluated antioxidant activity of compounds as a function of polarity in different lipid systems and reported that polar antioxidants are more effective in bulk oils, whereas nonpolar antioxidants are more effective in O/W emulsions. This hypothesis is known as the “antioxidant polar paradox”.4 Some researchers have also used this model to explain antioxidant behaviors in food systems. For example, this phenomenon can explain why polar antioxidants partition into © XXXX American Chemical Society
association colloids in bulk oil and are more effective than nonpolar antioxidants.5 In addition, nonpolar antioxidants are thought to be more effective in O/W emulsions because they are retained in the oil droplet.6 Further research has suggested that, in O/W emulsions, free radical scavengers are most effective when they concentrate at the oil−water interface of the emulsion droplets, where lipid oxidation reactions are most prevalent.7,8 Tocopherols are considered one of the most important nonpolar free radical scavenging antioxidants in foods and biological tissues. Tocopherols decrease the rate of lipid oxidation by donating hydrogen to a fatty acid radical and forming stable phenolic free radicals.9 Tocopherol homologues differ in polarity because of their number of methyl groups, with δ-, β-, γ-, and α-tocopherols containing 1, 2, 2, and 3 methyl groups, respectively (Figure 1). β- and γ-tocopherols both contain two methyl groups but in different positions.10 The amount of γ-tocopherol is greater than α- and δtocopherols in soybean oil, and β-tocopherol concentrations are very low.11 The tocopherol homologues have different antioxidant activity in O/W emulsions and bulk oils, which are Received: July 22, 2014 Revised: October 3, 2014 Accepted: October 9, 2014
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dx.doi.org/10.1021/jf503115j | J. Agric. Food Chem. XXXX, XXX, XXX−XXX
Journal of Agricultural and Food Chemistry
Article
Figure 1. Structures of tocopherol homologues used in this study. evaporated under a nitrogen stream. The SSO was flushed with nitrogen and stored at −80 °C until use. Removal of tocopherols in SSO was verified by HPLC. Emulsion Preparation. O/W emulsions were prepared using 1.0 wt % SSO in a 10 mM phosphate buffer solution (pH 7.0). Tween 20 was used as an emulsifier at a 1:10 emulsifier/oil ratio. SSO, Tween 20, and phosphate buffer were added to a beaker, and a coarse emulsion was made by blending with a hand-held two-speed homogenizer (model M133/1281-0, Biospec Products, Inc., Bartlesville, OK) at the high speed setting for 2 min. The coarse emulsion was then homogenized with a microfluidizer (model M-110L, Microfluidics, Newton, MA) at a pressure of 9 kbar for three passes. During homogenization, ice was used to cover the homogenizer chamber and coil, to maintain the emulsion temperature at γ- > α-tocopherols at all of the Tween 20 concentrations tested. At 1.0% Tween 20, the amount of α-, γ-, and δtocopherols in the aqueous phase was 73, 89, and 90%, respectively (Figure 2). δ-Tocopherol is the most polar and most surface-active20 of all of the tocopherols tested, which could explain why its solubilization into the aqueous phase by Tween 20 was greatest. Other researchers have reported that antioxidants in the emulsion droplet can migrate to the aqueous phase in the presence of surfactants. Richards et al.14 found that Brij 700 was able to solubilize propyl gallate and butylated hydroxyltoluene out of emulsion droplets, with the more polar propyl gallate partitioning into the aqueous phase to a greater extent. Laguerre et al.8 found that Brij 35 solubilized cholorogenic acid fatty acid esters out of emulsion droplets. Panya et al.15 reported that Tween 20 solubilized rosmarinic acid esters out of the oil droplets, but in this case, the more nonpolar 20 carbon ester partitioned into the aqueous phase more than the 4 carbon ester. It should be noted that the concentration of tocopherols in the presence of no added Tween 20 was in the order of δ- > γC
dx.doi.org/10.1021/jf503115j | J. Agric. Food Chem. XXXX, XXX, XXX−XXX
Journal of Agricultural and Food Chemistry
Article
Figure 3. Front-face fluorescence measurements (λex, 299; λem, 330) of 30 μM α-tocopherol in 1% stripped soybean O/W emulsions with varying Tween 20 concentrations. Data points and error bars represent means (n = 3) ± standard deviations.
Figure 4. (A) Lipid hydroperoxide and (B) hexanal formation in 1% SSO−Tween 20 emulsions in the absence (control) or presence of αtocopherol (35 μM) as a function of added Tween 20 (0, 0.1, 0.5, and 1.0%, w/w) during storage at 37 °C. Data points and error bars represent means (n = 3) ± standard deviations.
> α-tocopherols at 0.1% Tween 20. All three tocopherol homologues are listed in the Merck Index as not being soluble in water.21 During emulsion preparation, surfactants will concentrate at the droplet surface until the surface is saturated with the remaining surfactant partitioning into the aqueous phase.14 Tween 20 has a low critical micelle concentration (cmc;
