Lipid Oxidation in Oil-In-Water Emulsions
The Authors: Donpon Wannasin, Celina Fonseca and Eric A. Decker, Department of Food Science, University of Massachusetts, Amherst, USA
Many food products, for example, mayonnaise, salad dressing, and milk are emulsions. Emulsion contains two immersible phases where one phase is dispersed in the other phase as spherical droplets . Foods where oil droplets are dispersed in the aqueous phase are called oil-in-water (o/w) emulsions. Emulsions are stabilized by surface active emulsifiers that partition at the oil-water interface. Food emulsions are susceptible to lipid oxidation which causes formation of undesirable flavors and loss of nutrients . The properties of the oil-water interface are an important factor influencing lipid oxidation reaction as interfacial properties influences the location and reactivity of both prooxidants and antioxidants . This review will cover the mechanisms of lipid oxidation in oil-in water emulsions and methods to control oxidative deterioration.
2. Proxidants in oil-in water emulsions
Many prooxidants affect the mechanism of lipid oxidation in oil-in water emulsions. These prooxidants include transition metals, singlet oxygen, and location of hydroperoxides.
2.1 Role of transition metals in lipid oxidation
Transition metals are important prooxidants in oil-in-water emulsions. Both copper and iron promote lipid oxidation in emulsions. Transferrin, an iron specific chelator, strongly inhibited lipid oxidation in algae oil-in-water emulsions suggesting that iron is a more important prooxidant than copper . This is likely due to iron being at higher concentrations than copper in most food systems.
Iron and copper primarily promote oxidation by decomposing lipid hydroperoxides into free radicals. The addition of oxygen to fatty acid increases the polarity of the fatty acids which allows them to migrate to and concentrate at lipid droplet interfaces where they can rapidly react with aqueous phase metals or metals on the emulsion droplet surface. Reduced transition metals are more active at decomposing hydroperoxides. Transition metal can be reduced in foods by compounds such as ascorbic acid, carotenoids, plant phenolics and superoxide anion [5, 32].
2.2 Singlet Oxygen
Singlet oxygen is a major cause of lipid oxidation in oil-in-water emulsions in the presence of light. The primary photosensitizers in oil-in-water emulsions are chlorophyll and riboflavin. These photosensitizers can absorb light and become electronically excited and then transfer this energy to triplet oxygen to form singlet oxygen. The electron distribution of singlet oxygen allows it to directly add to the double bonds of unsaturated fatty acids to form hydroperoxides . Riboflavin can produce both singlet oxygen and superoxide anion in emulsions resulting in the formation of lipid hydroperoxides and the reduction of metals by superoxide anion to rapidly promote oxidation .
3. Factors that influence lipid oxidation in oil-in-water emulsions
3.1 Emulsion droplet size
Emulsification of lipids into droplets creates a large surface area for the interaction of prooxidants, free radicals and antioxidants at the emulsion droplet interface. Studies on how surface area influences oxidative stability are variable with a general conclusion being that surface area is not a major driver that changes oxidation rates . This could be because the large emulsion droplet surface area does not limit oxidation rates. In a study that used flow cytometry to study oxidation in individual droplets, it was found that smaller droplets oxidized slightly faster than larger droplets . However, oil-in-water emulsions typically contain a broad range of particle sizes so it may be experimentally difficult to observed changes in oxidation rates by changing the mean emulsion droplet size of an oil-in-water emulsion that has a multimodal participle size distribution.
3.2 Emulsion droplet interfacial properties
The properties of the interfacial layer of emulsion droplets is important because it can influence the ability of transition metals to interact with fatty acid hydroperoxide as well as impacting the ability of antioxidants to interact with free radicals . The charge of the emulsion droplet interface can impact lipid oxidation rates [4, 37]. Negatively charged emulsifiers were found to increase lipid oxidation as they attract cationic metals, whereas the positively charged emulsifiers can repel the cationic metals and slow down lipid oxidation [29, 30, 37]. Some studies have found a contradicting result where the negatively charged emulsifiers produce a lower lipid oxidation rate than the positively charged emulsifiers . This can be the result of excess negatively charged emulsifiers partitioning into the aqueous phase where they can bind and partition metals away from the emulsion droplet interface .
The thickness of the emulsion droplet interface can also impact oxidation rates. Silvestre et al. (2000) found that emulsifier head group size influences lipid oxidation rates with larger head groups inhibiting oxidation. However, tail group size was not found to impact lipid oxidation rates . Inhibition of oxidation by emulsifier head groups was thought to be due to inhibition of metal-hydroperoxide interactions.
Berton et. al (2012) compared oxidation rate between Tween 20-stabilized emulsion and protein-stabilized emulsion (β-lactoglobulin, β-bovineserum albumin, and β-casein) and found that protein-stabilized emulsion had a higher oxidation rate than Tween 20 stabilized emulsion both with and without initiators (iron plus ascorbic acid, metmyoglobin or 2,20-azobis(2-amidinopropane)-dihydrochloride). It was postulated that Tween 20 could form a more homogeneous and compact interfacial layer than protein thus decreasing prooxidant-emulsion droplet interactions.
3.3 pH and salt
The effects of pH on the oxidative stability of lipids in an oil-in-water emulsion found that oxidative stability decreased as pH increased . Waraho and coworkers (2011) found that the negative charge of Tween 20 stabilized oil-in-water emulsions decreased with increasing pH. This suggests that loss of negative charged could decrease metal-emulsion droplet interactions and be responsible for slower oxidation rates at higher pH. However, in protein stabilized emulsions, the charge of the emulsion droplet becomes positive when the pH is below the pI of the protein. These positively charged protein-stabilized emulsion droplets oxidize slower presumably due to their ability to repel transition metals and decrease metal-lipid interactions . Both sodium and potassium chloride increase oxidation rates in oil-in-water emulsions .
3.4 Oxygen concentration
Oxygen is a critical substrate in lipid oxidation reactions so its concentration and location can greatly influence oxidation rates. Oxygen is soluble in both lipid and water . Commercial emulsion formation is commonly done in open atmospheric conditions that expose all emulsion components to oxygen. This means that an emulsion would have oxygen in the lipid droplets, continuous phase and headspace. Marcuse and Fredricksson (1968) studied the effect of oxygen concentration on lipid oxidation in oil-in-water emulsions and found that the step where oxygen diffuse through aqueous phase to the lipid droplet was the rate-limiting step for lipid oxidation in the emulsions. At low oxygen concentration, mechanical agitation and cooling increased the lipid oxidation rate as they increased oxygen concentrations in the emulsion. However, at high oxygen concentration, these two processes had little impact on the lipid oxidation as the rate of oxygen diffusion was not limiting. Johnson and coworkers (2018) found that at least 58% of soluble oxygen had to be removed to decrease the rate of oxidation in a fish oil-in-water emulsion and 93% oxygen reduction was needed to get a substantial increase in oxidative stability [18, 19]. This study also showed that removal of headspace oxygen was not effective at decreasing lipid oxidation rates unless the emulsions was flushed with nitrogen for over 60 min suggesting that nitrogen flushing is not an effective way to stabilize oil-in-water emulsions.
4. Control of lipid oxidation in oil-in-water emulsions
4.1 Free radical scavenging antioxidants
Porter (1993) proposed the antioxidant polar paradox which stated that polar antioxidants work best in bulk oil while nonpolar antioxidants are most effective in oil-in-water emulsions. This hypothesis has been studied using free radical scavengers with similar radical scavenging activity that vary in polarity. The group of Edwin Frankel was initially active in this area using -tocopherol and Trolox with the non-polar -tocopherol being more effective in oil-in-water emulsions . This work was extended to other antioxidant pairs to further reinforce that nonpolar antioxidants are more effective in oil-in-water emulsions .
While the antioxidant polar paradox hypothesis states that non-polar antioxidant are more effective in oil-in-water emulsions, it was not able to determine if the surface activity of an antioxidant will impact its efficacy. Use of the interfacial probe, arenediazonium ion, was able to show that over 70% of -tocopherol partitioned at the interface of a Brij 20 stabilized oil-in-water emulsion , showing that antioxidants partition at the interface. The group of Villeneuve further advanced this field by creating surface active antioxidants by conjugating antioxidants to aliphatic chains of various carbon number to produce what was called phenolipids. This work showed that intermediate polarity antioxidants (8-12aliphatic chains) had the highest interfacial activity and antioxidant activity further supporting that antioxidants which concentrate at the emulsion droplet interface are the most effective .
Chelators are common additives used in slowing lipid oxidation caused by metals. EDTA is a strong chelator in oil-in-water emulsions whereas citrate and polyphosphates were ineffective in whey protein stabilized emulsions . EDTA is most effective when its concentration is greater than iron concentrations . Negatively charged, aqueous phase polysaccharides, proteins and peptides can also inhibit lipid oxidation by partitioning metals away from the emulsion droplet [7, 11].
5. Future ways to control lipid oxidation in oil-in-water emulsions
5.1 Multilayered interfaces
Since lipid oxidation reactions are predominately at the interface of oil-in-water emulsions, modifying the emulsion droplet surface can be an effective way to inhibit oxidative deterioration. Multilayered emulsion droplet interfaces can be created by layer-by-layer deposition techniques where charged polymer is absorbed onto an oppositely charged emulsion droplet. These multilayer emulsion droplets can increase oxidative stability by creating thick interfacial layers and/or by creating positive charges that repel iron . It is difficult to make multilayer emulsions with high lipid concentrations which have limited their commercial applications.
5.2 Active packaging
Active packaging is another strategy that can be used to prevent lipid oxidation. Many active substances, for example, metal chelators, free radical scavengers, and oxygen scavengers, can be added to the food contact layer of the packaging to prevent oxidation and extend the shelf life of food products . Tian and coworkers developed metal chelating packaging material that was able to inhibit lipid oxidation in oil-in-water emulsions  but these films are not currently available. Active packaging material that absorbs oxygen can also be used to control lipid oxidation. Johnson and coworkers (2018) found that an oxygen absorbing film developed by Mitsubishi was very effective at inhibiting lipid oxidation in oil-in-water emulsions.
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