Effects of salts on oxidative stability of lipids in Tween-20 stabilized oil-in-water emulsions

Accepted Manuscript Effects of salts on oxidative stability of lipids in tween-20 stabilized oil-in-water emulsions Leqi Cui, Hyung Taek Cho, D. Julian McClements, Eric A. Decker, Yeonhwa Park PII: DOI: Reference:

S0308-8146(15)30244-2 http://dx.doi.org/10.1016/j.foodchem.2015.11.099 FOCH 18440

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Food Chemistry

Received Date: Revised Date: Accepted Date:

2 April 2015 8 October 2015 19 November 2015

Please cite this article as: Cui, L., Cho, H.T., Julian McClements, D., Decker, E.A., Park, Y., Effects of salts on oxidative stability of lipids in tween-20 stabilized oil-in-water emulsions, Food Chemistry (2015), doi: http:// dx.doi.org/10.1016/j.foodchem.2015.11.099

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Effects of salts on oxidative stability of lipids in tween-20 stabilized oil-in-water emulsions

Leqi Cui1, Hyung Taek Cho1, D. Julian McClements1,2, Eric A. Decker1,2 and Yeonhwa Park1*

1

Department of Food Science, University of Massachusetts,

102 Holdsworth Way, Amherst, Massachusetts 01003, USA

2

Department of Biochemistry, Faculty of Science, King Abdulaziz University, P. O. Box 80203 Jeddah 21589 Saudi Arabia

*

To whom correspondence should be addressed

Department of Food Science, University of Massachusetts 102 Holdsworth Way, Amherst, MA 01003 Telephone: (413) 545-1018, Fax: (413) 545-1262; e-mail: [email protected]

1

Abstract Lipid oxidation in oil-in-water (O/W) emulsions is an important factor determining the shelf life of food products. Salts are often present in many types of emulsion based food products. However, there is limited information on influence of salts on lipid oxidation in O/W emulsions. Thus, the purpose of this study was to examine the effects of sodium and potassium chloride on lipid oxidation in O/W emulsions. Tween 20 stabilized corn O/W emulsions at pH 7.0 were prepared with different concentrations of sodium chloride with or without the metal chelators. NaCl did not cause any changes in emulsion droplet size. NaCl dose-dependently promoted lipid oxidation as measured by the lipid oxidation product, hexanal. Both deferoxamine (DFO) and ethylenediaminetetraacetic acid (EDTA) reduced lipid oxidation in emulsions with NaCl, with EDTA being more effective. Potassium chloride showed similar impact on lipid oxidation as sodium chloride. These results suggest that salts are able to promote lipid oxidation in emulsions and this effect can be controlled by metal chelators.

Keywords: salts; emulsion; lipid oxidation; iron; chelator

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1. Introduction Lipid oxidation is one of the major contributing factors on the quality and shelf life of natural and processed foods (Frankel, 2005). It adversely influences flavor as well as quality by forming a number of oxidative products from lipid, such as lipid hydroperoxides, polymeric materials and fatty acid chain-cleavage products (Shao & Heinecke, 2009; Waraho, McClements, & Decker, 2011). Often lipids in foods are present as O/W emulsions, and oxidation reactions of lipids are influenced by various factors including prooxidants, antioxidants, increased surface area, emulsion droplet interface characteristics and the presence of metal ions (Berton-Carabin, Ropers, & Genot, 2014; McClements & Decker, 2000; Waraho, McClements, & Decker, 2011). The lipid emulsions usually contain small quantities of lipid peroxides (Mancuso, McClements, & Decker, 1999) which can be decomposed by transition metal ions originated from water and food ingredients into free radicals that promote oxidation (Dunford, 1987). Salt, most often sodium chloride, is commonly used in food, cosmetics, and pharmaceutical formulations. Sodium chloride in food plays a significant role in taste by way of contributing to flavor, texture and aroma, as well as playing an important role in food safety by contributing to the prevention of microbial growth (Cervantes, Lund, & Olson, 1983; Floury, Camier, Rousseau, Lopez, Tissier, & Famelart, 2009; Poll & Flink, 1984; Taormina, 2010). Many foods exist partly or wholly as an emulsion with salt, such as milk, butter, cream, soups, margarine, sauces and mayonnaise (Dickinson & Stainsby, 1982; Friberg, Larsson, & Sjoblom, 2003). More specifically, peanut butter, margarine-butter blend, ice cream, beverage (protein powder soy based) and mayonnaise usually contain 0.54, 0.72, 0.16, 0.16, 0.88 wt% (ranging from 70 mmol/kg to 380 mmol/kg) sodium respectively (U.S. Department of Agriculture, 2013). The effect of salts on the oxidation of commercial soybean oil (Calligaris & Nicoli, 2006) and

3

ascorbic acid (Harel, 1994) has been reported before. However, there is limited information on the role of sodium and potassium chloride in lipid oxidation in O/W emulsion. Shao and Tang (Shao & Tang, 2014) reported antioxidant effect of 300 mM sodium chloride on a soy protein isolate (SPI)-stabilized soybean oil-in-water emulsion. Our lab previously reported both prooxidative and no impact of sodium chloride activity in lipid oxidation in O/W emulsion with different emulsifiers. However, these emulsions had added iron at concentrations greater than seen in typical food emulsions (Mei, Decker, & McClements, 1998; Mei, McClements, Wu, & Decker, 1998; Shao & Tang, 2014). NaCl was found to be prooxidative in oil-in-water emulsions oxidized by copper at pH 3.0 (Osborn-Barnes & Akoh, 2003). Arnold and coworkers (Arnold, Bascetta, & Gunstone, 1991) found that in pork phospholipid liposomes, sodium chloride inhibited the prooxidative effect of copper at 4, but enhanced its ability at -8 and -20°C. Kristinová and coworkers (Kristinová, Mozuraityte, Storrø, & Rustad, 2009) examined the impact of different salts on the oxidation of phospholipid liposome. By measuring oxygen uptake, they found that cations and some anions (sulphates and nitrates) did not influence the oxidation rates, while some anions (chloride and phosphate) reduced oxidation rates. Oxidation in liposomes could be different from oil-in-water emulsions since their physical structures would be different and phospholipids have been shown to be both antioxidative and prooxidative (Cui, Kittipongpittaya, McClements, & Decker, 2014; Takenaka, Hosokawa, & Miyashita, 2007). In addition, all of these studies added high levels of transition metals to promote oxidation and thus might not reflect how NaCl impacts oxidation in food emulsions. To get a better understanding of how sodium chloride impacts lipid oxidation in O/W emulsion containing endogenous iron concentrations, the nonionic surfactant Tween 20 was used to stabilize corn O/W emulsions at

4

pH 7.0 and the role of salts with or without metal chelators on lipid oxidation in this model system was investigated.

2. Experimental 2.1 Materials All reagents were of analytical grade. Corn oil was purchased from a local store and stored at 4 °C in dark. Tween 20, deferoxamine (DFO), dipotassium phosphate, monopotassium phosphate, potassium chloride, ethylenediaminetetraacetic acid (EDTA) and hexanal were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). Sodium chloride (contains less than 0.01 wt% Bromide, 0.002 wt% Calcium, 0.001 wt% Magnesium, 0.005 wt% Potassium, 5 ppm Lead and 2 ppm Iron) was purchased from Fisher Scientific Chemical Co. (Pittsburgh, PA). Distilled and deionized water was used in all experiments.

2.2 Preparation of emulsions The aqueous phase of the emulsion was prepared by dispersing 1.0 wt% Tween 20 in 5 mM phosphate buffer at pH 7.0 without and with DFO (50 µM) or EDTA (50 µM) followed by stirring at room temperature overnight to ensure complete dispersion and hydration. Corn oil-inwater emulsions were prepared by homogenizing 10 wt% oil phase with 90 wt% aqueous phase at ambient temperature. A coarse emulsion was prepared using a high-speed blender for 2 min, (Biospec Products Inc., Bartlesville, USA), which was then passed through a high pressure homogenizer (Model 101, Microfluidics, Newton, Massachusetts, USA) three times at 9000 psi. During each pass, the emulsions were collected in a beaker submerged in a cool water bath. Sodium chloride at 0.6, 1.0 and 1.6 wt% (of the total emulsion) and potassium chloride at 100

5

mmol/kg (same molar concentration as 0.6 wt % sodium chloride) were then added to samples of emulsions. Samples (1 mL) were placed in 10-mL headspace vials sealed with PTFE/silicone septa lined aluminum caps and stored at 37 °C in the dark.

2.3 Evaluation of physical properties of emulsions The particle size distribution (PSD) of all samples was measured by static light scattering using a commercial instrument (Mastersizer 2000), with a small volume sample dispersion unit (Hydro 2000 SM) (Malvern, Worcestershire, UK). All samples were diluted in 5 mM phosphate buffer adjusted to pH 7.0, and the stir speed for the small volume sample dispersion unit was set to 1250 rpm. The refractive indexes used for particle and dispersant phase are 1.474 and 1.33 respectively. Each sample was measured in duplicate. The ζ-potential of all samples was measured by laser Doppler electrophoresis (Zetamaster, Malvern, Worcestershire, UK). Prior to measuring, samples were diluted with 5 mM phosphate buffer at pH 7.0 to a droplet concentration of approximately 0.001 wt%. Five measurements were taken per sample injected, and each sample was measured in duplicate for a total of ten ζ-potential readings per sample.

2.4 Measurement of Oxidation Parameters. Hexanal concentration was measured using a GC-17A Shimadzu gas chromatograph equipped with an AOC-5000 autosampler (Shimadzu, Kyoto, Japan). A 30 m × 0.32 mm Equity DB-1 column (Supelco, Bellefonte, PA) with a 1 µm film thickness was used for separations. Each sample was heated at 55°C in the autosampler heating block for 15 min. A 50/30 µm DVB/ Carboxen/ PDMS solid-phase microextraction (SPME) fiber needle (Supelco) was injected into the sample vial headspace for 2 min to adsorb volatiles and then was inserted into the 250°C

6

injector port for 3 min at a split ratio of 1:5. The gas chromatograph ran for 20 min at 65°C for each sample. Helium was used as a carrier gas, with a total flow rate of 15.0 mL/min. A flame ionization detector at a temperature of 250°C was used. Peak integration was performed using Shimadzu Class-VP (version 7.4). Concentrations were calculated by using a standard curve made from the fresh emulsions containing known hexanal concentrations and individual salt concentrations.

2.5 Statistical analysis All data were obtained from triplicate samples or more and expressed as the mean ± standard deviation. Statistical differences of the experimental results were determined by analysis of variance (ANOVA) in the SAS statistical software package (SAS Inst. Inc., Cary, NC). Duncan’s multiple-range test was used to determine differences between means, and p<0.05 was considered to be statistically significant.

3. Results and Discussion 3.1 Impact of salt concentrations on the physical properties of oil in water emulsions Corn O/W emulsions were prepared and stabilized by nonionic surfactant Tween 20. The effects of salt concentrations on the physical properties of emulsions was determined and the particle size and zeta potential of emulsions with four levels of salt in absence or presence of metal chelator DFO are shown in Table 1. Zeta potential was measured since changes in droplet charge could impact lipid oxidation rates. All emulsions had similar d43 particle size of around 0.31µm, showing that neither sodium chloride nor its combination with DFO changed particle size of these emulsions. These droplet sizes are similar to those found in many emulsion-based

7

food products such as soft drinks and cream liqueurs (McClements, 2005). As for zeta potential of emulsions in the absence of DFO, control emulsion had a -3.5 mV surface charge as compared to -2.9, -2.8 and -2.7 mV of emulsions containing 0.6, 1.0, 1.6 wt% sodium chloride respectively. These differences were not statistically significant. Although the Tween 20 used in this experiment is often considered nonionic, it was not surprising to see a negative emulsion surface charge since other researchers have also reported this phenomenon (Hur, Decker, & McClements, 2009; Waraho, Cardenia, Rodriguez-Estrada, McClements, & Decker, 2009). The reason for this negative surface charge could be due to either impurities in the Tween 20 or free fatty acids in the system. No significant difference in zeta potential was seen in the presence of DFO and sodium chloride with all four groups being -3.0 or -3.1 mV. Monovalent ions generally do not change the charge of emulsion droplets (D. McClements, 2005). No changes in zeta potential and droplet size and no visual creaming were observed during the storage studies.

3.2 Impact of sodium chloride concentrations on the chemical properties of oil in water emulsions The effect of sodium chloride concentrations on the oxidative stability of emulsions was determined by monitoring the formation of hexanal, a secondary lipid oxidation product commonly produced from omega-6 fatty acids (Figure 1). As we can see from Figure 1, as sodium chloride concentration increased, the initial formation of hexanal was significantly accelerated and the lag phase of emulsion was shortened. For example, the control had a lag phase of 8 days compared to emulsions with 0.6 wt% sodium chloride whose lag phase was only 1-2 days. Emulsions with 1.0 and 1.6 wt% sodium chloride had no measurable oxidation lag phase.

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3.3 Impact of deferoxamine on salt associated lipid oxidation in oil in water emulsions Transition metals are well known to be one of the most important prooxidants in O/W emulsions (Frankel, 2005; McClements & Decker, 2000). Trace amount of metals in food products originate from water, food ingredients or food processing equipment (Reilly, 2008). Even in laboratory settings using double distilled water, high purity chemicals and acid washed glassware, there are enough contaminating metals to promote lipid oxidation. To determine if iron played an important role in lipid oxidation in our system, deferoxamine (DFO), a metal chelator, was added to the emulsions in combinations with different concentrations of sodium chloride (Figure 2). DFO reduced hexanal formation at all sodium chloride concentrations tested (Figure 2a-d). It is apparent that the addition of DFO did not completely block lipid oxidation in the O/W emulsions, suggesting that there could still be some prooxidative metals that do not interact strongly with DFO. The DFO-iron complex binding constant is 1031 while the DFO-copper binding constant is ≤ 1014 (Neilands, 1981). This compares to EDTA that has a higher copper binding constant of 1018 but lower iron binding constant of 1024 (Smith & Martell, 1989). EDTA was found to be more effective than DFO at inhibiting lipid oxidation in oil-in-water emulsions containing 0.6 wt% sodium chloride with the EDTA samples still being in the lag phase after 11 days of storage (Figure 3). These results suggest that copper may also be an important prooxidant in the salt promoted oxidation. However, this could also be due to the ability of DFO-iron complexes to weakly promote hydrogen peroxide decomposition into free radicals (Schaich & Borg, 1988). EDTA can also increase the prooxidant activity of iron but this occurs mainly at EDTA: iron ratios that are much lower (≤ 1) than used in this study (Mahoney & Graf, 1986).

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The observation that salt promotes lipid oxidation in O/W emulsions is in contrast to Mei et al. (Mei, McClements, Wu, & Decker, 1998) who found that sodium chloride had no impact on the formation of lipid hydroperoxides and thiobarbituric acid reactive substances in Brij 35 stabilized corn O/W emulsions where iron and ascorbic acid had been added to accelerate lipid oxidation and the corn oil had been stripped of polar compounds including tocopherols. In these emulsions, the Brij 35 produced similar zeta potential of approximately -3.0 mV (Mei, McClements, & Decker, 1999), so it is unlikely that emulsion droplet charge was responsible for the observed differences. The lack of prooxidative activity of sodium chloride reported by Mei et al. (Mei, McClements, Wu, & Decker, 1998) could be due to their use of a 50 µM iron and 150 µM ascorbate redox cycling system, which was designed to increase the prooxidant activity of iron by ascorbate reducing the iron to its more reactive ferrous state. However, if the prooxidant effect of sodium chloride was due to its ability to accelerate the prooxidant activity of iron, this redox cycling system could alter iron-sodium chloride interactions. In addition, the high level of iron used might eliminate the lag phase of oxidation, which could overwhelm the ability of NaCl to alter oxidation kinetics. Also, the prooxidant activity of sodium chloride might only be significant at low iron concentrations. An additional difference between this research and the previous study is the presence of tocopherols. Mei et al. (Mei, McClements, Wu, & Decker, 1998) used corn oil stripped of tocopherols whereas this research used a commercial grade refined oil. Recent work by Chen et al. (Chen, Panya, McClements, & Decker, 2012) found that iron is able to degrade tocopherols. In this study, they found that in a medium-chain triacylglycerides (MCT) bulk oil system which did not contain oxidizable unsaturated fatty acids and thus would not have free radicals generated by fatty acid oxidation, α-tocopherol reduced ferric iron thus causing a loss of α-tocopherol. If sodium chloride accelerated iron-promoted

10

tocopherol degradation in the emulsions used in our research, this could further explain why sodium chloride was more prooxidative than in the work of Mei et al. (Mei, McClements, Wu, & Decker, 1998).

3.4 Comparison between sodium chloride and potassium chloride on lipid oxidation in oil in water emulsions The prooxidant activity of sodium chloride has been postulated to be due to the ability of sodium to displace iron from macromolecules such as proteins (Kanner, Harel, & Jaffe, 1991) or the ability of chloride to form prooxidative complexes with transition metals (Osinchak, Hultin, Zajicek, Kelleher, & Huang, 1992). The former mechanism (Kanner, Harel, & Jaffe, 1991) was evaluated by measuring how much chelatable iron from muscle could be extracted by distilled water and 0.3 M NaCl. It was found that NaCl increased the extraction of iron ions from muscle tissue, suggesting that sodium chloride increased the removal of iron from membranes or proteins by sodium displacement or chloride forming complexes with iron. The latter mechanism (Osinchak, Hultin, Zajicek, Kelleher, & Huang, 1992) was evaluated by examining the impact of different cations and anions on the oxidation of a liposome model system containing mackerel muscle press juice. The authors found Cl being the active prooxidant in their system. To test if the observed prooxidant activity of sodium chloride in our O/W emulsions was due to the sodium, we compared the prooxidant activity of sodium to potassium chloride at the same concentration (100 mM) (Figure 4). A new batch of oil was used for these experiments which could explain why the lag phase of the control was longer. However, this experiment was consistent with previous experiments since the sodium chloride again decreased the hexanal lag phase by 5-6 days. The KCl and NaCl produced similar oxidation kinetics indicating that

11

potassium chloride could also promote lipid oxidation in O/W emulsions. This suggested the cations (sodium and potassium) were not responsible for the observed prooxidant activity of these salts in the O/W emulsion. One concern about the prooxidant activity of salts shown in this study could be the trace amount of metals that the chemicals contained. According to the manufacturers, the salts used here contained iron (≤2 ppm in NaCl and ≤5 ppm in KCl) as well as heavy metals such as Pb (≤5 ppm in NaCl and ≤1 ppm in KCl). We tested the impact of 0.012, 0.02 and 0.032 ppm ferric chloride on the oxidative stability of the emulsions as these were the maximal iron concentrations that could be contributed by 0.6, 1.0 and 1.6 wt% sodium chloride. The results showed that these levels of iron did not accelerate oxidation as there were no differences in the oxidation lag phases (data now shown). Osinchak et al. (Osinchak, Hultin, Zajicek, Kelleher, & Huang, 1992) also reported that 0.1 ppm iron had little effect on oxidation rates in liposomes (<0.1% ).

4. Conclusions Overall our current results suggest the prooxidant property of salt in O/W emulsions is linked to its ability to increase the activity of naturally occurring metal ions since metal chelators can inhibit salt promoted lipid oxidation in O/W emulsions. This suggest that salt-promoted lipid oxidation could be significant in many food emulsions as products such as mayonnaise and butter which have similar sodium chloride concentrations (U.S. Department of Agriculture, 2013) as used in this study. In addition to sodium chloride increasing the activity of metals, it could also be a source of prooxidant metals. While the NaCl used in this study contained less

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than 2 ppm iron, commercial sources have been reported to contain up to 18 ppm iron suggesting that contaminating metals in salt could further promote oxidation (Heshmati, Vahidinia, & Salehi, 2014; Khaniki, Dehghani, Mahvi, & Nazmara, 2007). This work also suggests that substituting sodium chloride with potassium chloride will not be a solution to decrease NaCl-promoted oxidation. Instead, strategies to replace chloride might be more successful. Finally, additional work using sensory analysis would be useful in gaining a more complete understanding of how salts impact lipid oxidation. While hexanal is a useful marker of lipid oxidation and was able to show the prooxidant activity of salt, NaCl could also change sensory perception by increasing volatility of lipid oxidation breakdown products and/or masking or enhancing rancid off-flavors.

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Reference Arnold, A. R., Bascetta, E., & Gunstone, F. D. (1991). Effect of Sodium Chloride on Pro-oxidant Activity of Copper (II) in Peroxidation of Phospholipid Liposomes. Journal Of Food Science, 56(2), 571-573. Berton-Carabin, C. C., Ropers, M.-H., & Genot, C. (2014). Lipid Oxidation in Oil-in-Water Emulsions: Involvement of the Interfacial Layer. Comprehensive Reviews in Food Science and Food Safety, 13(5), 945-977. Calligaris, S., & Nicoli, M. C. (2006). Effect of selected ions from lyotropic series on lipid oxidation rate. Food Chemistry, 94(1), 130-134. Cervantes, M. A., Lund, D. B., & Olson, N. F. (1983). Effects of salt concentration and freezing on Mozzarella cheese texture. Journal of Dairy Science, 66(2), 204-213. Chen, B., Panya, A., McClements, D. J., & Decker, E. A. (2012). New Insights into the Role of Iron in the Promotion of Lipid Oxidation in Bulk Oils Containing Reverse Micelles. Journal Of Agricultural and Food Chemistry, 60(13), 3524-3532. Cui, L., Kittipongpittaya, K., McClements, D. J., & Decker, E. (2014). Impact of Phosphoethanolamine Reverse Micelles on Lipid Oxidation in Bulk Oils. Journal of the American Oil Chemists' Society, 91(11), 1931-1937. Dickinson, E., & Stainsby, G. (1982). Colloids in food: Applied Science Publishers. Dunford, H. B. (1987). Free radicals in iron-containing systems. Free Radical Biology and Medicine, 3(6), 405-421. Floury, J., Camier, B., Rousseau, F., Lopez, C., Tissier, J.-P., & Famelart, M.-H. (2009). Reducing salt level in food: Part 1. Factors affecting the manufacture of model cheese systems and their structure– texture relationships. LWT-Food Science and Technology, 42(10), 1611-1620. Frankel, E. N. (2005). Lipid oxidation: The Oily Press. Friberg, S., Larsson, K., & Sjoblom, J. (2003). Food emulsions: CRC Press. Harel, S. (1994). Oxidation of ascorbic acid and metal ions as affected by NaCl. Journal Of Agricultural and Food Chemistry, 42(11), 2402-2406. Heshmati, A., Vahidinia, A., & Salehi, I. (2014). Determination of Heavy Metal Levels in Edible Salt. Avicenna J Med Biochem, 2(1), e19836. Hur, S. J., Decker, E. A., & McClements, D. J. (2009). Influence of initial emulsifier type on microstructural changes occurring in emulsified lipids during in vitro digestion. Food Chemistry, 114(1), 253-262. Kanner, J., Harel, S., & Jaffe, R. (1991). Lipid peroxidation of muscle food as affected by sodium chloride. Journal Of Agricultural and Food Chemistry, 39(6), 1017-1021. Khaniki, G. R. J., Dehghani, M. H., Mahvi, A. H., & Nazmara, S. (2007). Determination of trace metal contaminants in edible salts in Tehran (Iran) by atomic absorption spectrophotometry. J Biol Sci, 7(5), 811-814. Kristinová, V., Mozuraityte, R., Storrø, I., & Rustad, T. (2009). Antioxidant Activity of Phenolic Acids in Lipid Oxidation Catalyzed by Different Prooxidants. Journal Of Agricultural and Food Chemistry, 57(21), 10377-10385. Mahoney, J. R., & Graf, E. (1986). Role of Alpha-Tocopherol, Ascorbic Acid, Citric Acidand EDTA as Oxidants in Model Systems. Journal Of Food Science, 51(5), 1293-1296. Mancuso, J. R., McClements, D. J., & Decker, E. A. (1999). Ability of Iron To Promote Surfactant Peroxide Decomposition and Oxidize α-Tocopherol. Journal Of Agricultural and Food Chemistry, 47(10), 4146-4149. McClements, D. (2005). Food emulsions: principles, practices, and techniques. CRC series in contemporary food science.

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McClements, D. J., & Decker, E. A. (2000). Lipid Oxidation in Oil-in-Water Emulsions: Impact of Molecular Environment on Chemical Reactions in Heterogeneous Food Systems. Journal Of Food Science, 65(8), 1270-1282. Mei, L., Decker, E. A., & McClements, D. J. (1998). Evidence of Iron Association with Emulsion Droplets and Its Impact on Lipid Oxidation. Journal Of Agricultural and Food Chemistry, 46(12), 5072-5077. Mei, L., McClements, D. J., & Decker, E. A. (1999). Lipid Oxidation in Emulsions As Affected by Charge Status of Antioxidants and Emulsion Droplets. Journal Of Agricultural and Food Chemistry, 47(6), 2267-2273. Mei, L., McClements, D. J., Wu, J., & Decker, E. A. (1998). Iron-catalyzed lipid oxidation in emulsion as affected by surfactant, pH and NaCl. Food Chemistry, 61(3), 307-312. Neilands, J. B. (1981). Microbial Iron Compounds. Annual Review Of Biochemistry, 50(1), 715-731. Osborn-Barnes, H. T., & Akoh, C. C. (2003). Copper-Catalyzed Oxidation of a Structured Lipid-Based Emulsion Containing α-Tocopherol and Citric Acid:  Influence of pH and NaCl. Journal Of Agricultural and Food Chemistry, 51(23), 6851-6855. Osinchak, J. E., Hultin, H. O., Zajicek, O. T., Kelleher, S. D., & Huang, C. H. (1992). Effect of NaCl on catalysis of lipid oxidation by the soluble fraction of fish muscle. Free Radic Biol Med, 12(1), 3541. Poll, L., & Flink, J. M. (1984). Aroma analysis of apple juice: Influence of salt addition on headspace volatile composition as measured by gas chromatography and corresponding sensory evaluations. Food Chemistry, 13(3), 193-207. Reilly, C. (2008). Metal contamination of food: its significance for food quality and human health: John Wiley & Sons. Schaich, K. M., & Borg, D. C. (1988). Fenton reactions in lipid phases. Lipids, 23(6), 570-579. Shao, B., & Heinecke, J. W. (2009). HDL, lipid peroxidation, and atherosclerosis. Journal of lipid research, 50(4), 599-601. Shao, Y., & Tang, C.-H. (2014). Characteristics and oxidative stability of soy protein-stabilized oil-in-water emulsions: Influence of ionic strength and heat pretreatment. Food Hydrocolloids, 37, 149-158. Smith, R., & Martell, A. (1989). Aminocarboxylic Acids. In Critical Stability Constants, vol. 6 (pp. 1-66): Springer US. Takenaka, A., Hosokawa, M., & Miyashita, K. (2007). Unsaturated Phosphatidylethanolamine as Effective Synergist in Combination with α-Tocopherol. Journal of Oleo Science, 56(10), 511-516. Taormina, P. J. (2010). Implications of salt and sodium reduction on microbial food safety. Critical reviews in food science and nutrition, 50(3), 209-227. U.S. Department of Agriculture, A. R. S. (2013). USDA National Nutrient Database for Standard Reference, Release 26. Nutrient Data Laboratory Home Page, http://www.ars.usda.gov/ba/bhnrc/ndl. Waraho, T., Cardenia, V., Rodriguez-Estrada, M. T., McClements, D. J., & Decker, E. A. (2009). Prooxidant mechanisms of free fatty acids in stripped soybean oil-in-water emulsions. J Agric Food Chem, 57(15), 7112-7117. Waraho, T., McClements, D. J., & Decker, E. A. (2011). Mechanisms of lipid oxidation in food dispersions. Trends in Food Science & Technology, 22(1), 3-13.

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Figure Captions

Table 1. Influence of sodium chloride and deferoxamine on the electrical charge and diameter of emulsion droplets.

Figure 1. Effects of sodium chloride on hexanal production in 10 wt% corn oil-in-water emulsions. Emulsions were incubated at 37 ℃. Data are means ± standard deviations (n = 3). Means with different letters are significantly different at P<0.05 at each time point.

Figure 2. Effects of deferoxamine on hexanal production in various concentrations of sodium chloride in 10 wt% corn oil-in-water emulsions. 50 µM deferoxamine were used without (A) or with 0.6 (B), 1.0 (C) and 1.6 wt% (D) sodium chloride at 37 ℃. Data are means ± standard deviations (n = 3). *P<0.05 as compare to no deferoxamine containing emulsion at each time point.

Figure 3. Comparison of deferoxamine (50 µM) and EDTA (50 µM) on hexanal formation in 10 wt% corn oil-in-water emulsions containing 0.6 wt% sodium chloride. Emulsions were incubated at 37 ℃. Data are means ± standard deviations (n = 3). *P<0.05 as compare to control. #P < 0.05 as compare to deferoxamine containing emulsions.

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Figure 4. Comparison of sodium chloride and potassium chloride on hexanal formation in corn oil-in-water emulsions. Both were 100 mM and incubated at 37 ℃. Data are means ± standard deviations (n = 3).

Table 1. Emulsion without deferoxamine

Emulsion with deferoxamine

Concentration of sodium chloride (%)

d43 (µm)

ζ –potential (mV)

d43 (µm)

ζ –potential (mV)

0

0.31 ± 0.01

-3.5 ± 0.2

0.31 ± 0.01

-3.0 ± 0.3

0.6

0.32 ± 0.01

-2.8 ± 0.3

0.31 ± 0.01

-3.1 ± 0.2

1.0

0.32 ± 0.01

-2.9 ± 0.3

0.31 ± 0.01

-3.1 ± 0.2

1.6

0.31 ± 0.01

-2.7 ± 0.4

0.31 ± 0.01

-3.0 ± 0.3

17

12

Hexanal concentration (mmol/g oil)

a 0.0% 0.6% 1.0% 1.6%

10

a

a a

b a

8

a

6

b

a

c

a

4

a

a

b

a b

2

c

b d

0 0

b 2

b

d

b

c

4

6

8

Time (day)

10

12

A

Control Deferoxamine

10

Hexanal concentration (mmol/g oil)

Hexanal concentration (mmol/g oil)

12

8

6

4

2

B

Control Deferoxamine

10

8

6

*

4

* 2

* 0 0

2

4

6

*

8

*

0

*

0

10

2

4

Control Deferoxamine

8

6

* 4

* 2

*

0 0

8

12

C Hexanal concentration (mmol/g oil)

Hexanal concentration (mmol/g oil)

12

10

6

10

Time (day)

Time (day)

D Control Deferoxamine

10

8

6

*

4

*

2

* 0

2

4

6

Time (day)

8

10

0

2

4

6

Time (day)

8

10

Hexanal concentration (mmol/g oil)

5

Control Deferoxamine EDTA

4

*

3

2

1

0 0

, **

2

*,#

*

, ** 4

6

Time (day)

8

*,# 10

Hexanal concentration (mmol/ g oil)

12

Control NaCl KCl

10

8

6

4

2

0 0

5

10

15

Time (Day)

20

25

30

• • •

NaCl promoted lipid oxidation of tween-20 stabilized oil-in-water emulsions. Metal chelators DFO and EDTA were found to inhibit this effect. KCl produced similar lipid oxidation kinetics as NaCl in oil-in-water emulsions.

18