Article pubs.acs.org/JAFC
Synergy of Licorice Extract and Pea Protein Hydrolysate for Oxidative Stability of Soybean Oil-in-Water Emulsions Xin Zhang,† Youling L. Xiong,*,‡ Jie Chen,† and Lirong Zhou§ †
State Key Laboratory of Food Science and Technology and School of Food Science and Technology, Jiangnan University, Wuxi, Jiangsu 214122, China ‡ Department of Animal and Food Sciences, University of Kentucky, Lexington, Kentucky 40546, United States § ICL Performance Products LP, St. Louis, Missouri 63141, United States ABSTRACT: Previously developed radical-scavenging pea protein hydrolysates (PPHs) prepared with Flavourzyme (Fla-PPH) and Protamex (Pro-PPH) were used as cosurfactants with Tween 20 to produce soybean oil-in-water (O/W) emulsions, and the suppression of lipid oxidation was investigated. Both PPHs significantly retarded oxidation (P < 0.05) of the emulsions when stored at 37 °C for 14 days. Electron microscopy revealed an interfacial peptidyl membrane around oil droplets, which afforded steric restrictions to oxidation initiators. When licorice extract (LE) was also used in emulsion preparation, a remarkable synergistic oxidation inhibition was observed with both PPHs. LE adsorbed onto oil droplets either directly or through associating with PPH to produce a thick and compact interfacial membrane enabling the defense against oxygen species. Liquiritin apioside, neolicuroside, glabrene, and 18β-glycyrrhetic acid were the predominant phenolic derivatives partitioning at the interface and most likely the major contributors to the notable synergistic antioxidant activity when coupled with PPHs. KEYWORDS: antioxidant, pea protein hydrolysate, licorice extract, emulsion, oxidative stability, synergism
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INTRODUCTION A wide array of natural and processed foods exist as emulsions, such as milk, cream, mayonnaise, and salad dressings. Due to the large contact surface between oxidizable fatty acids and water-soluble oxidants (oxygen and transition metal ions), emulsified oil is much more susceptible to oxidation than bulk oil.1 Peptides and protein hydrolysates as potential alternatives to traditional antioxidants have been widely reported to be inhibitory of lipid oxidation in biphasic (O/W) systems through scavenging free radicals, sequestrating pro-oxidative metal ions, or forming physical (shielding) barrier around oil droplets.2−4 Unlike small-molecule antioxidants, protein hydrolysates may also act as stabilizing agents to improve the physical stability of emulsion systems.5,6 Phenolic compounds extracted from plants and spices are also recognized for their strong antioxidant activity in food systems due to radical-scavenging activity and chelation of transition metal ions.7,8 Many polyphenols and their plant extracts are effective against both lipid peroxidation and protein oxidative changes in model systems and in complex food matrices, such as processed meats.9,10 The effectiveness of an antioxidant in retarding lipid oxidation in food emulsions depends not only on its chemical reactivity with oxidation initiators and propagators but also on its location, which is dictated by its polarity and affinity for other constituents.11,12 Due to the size reduction and increased charges, peptides are distributed preferentially in the aqueous phase and can partition at the oil−water interface, thereby preventing the initiation of lipid oxidation induced by watersoluble oxidants. On the other hand, as polyhydroxyl phenol derivatives with both polar and nonpolar moieties, phenolic compounds have the propensity to concentrate at the oil−water interface and are dispersible in the lipid phase (where © 2014 American Chemical Society
hydroperoxides and peroxide radicals are generated) to curtail the propagation of lipid radical chain reactions. Because of their complementary processes in inhibiting lipid oxidation, peptides (protein) and phenolic compounds when used in combination have been found to function synergistically in several biphasic systems, such as O/W emulsions7,13,14 and a liposome suspension.15 However, it should be noted that similar to Tween, Span, and monoglycerides, phenolic molecules as surface active compounds might influence the adsorption of peptides at the interface either indirectly by competing for the oil droplets or directly by binding to peptides through hydrophobic interaction and hydrogen bonds.16 Therefore, the existence of phenolics in a peptide-stabilized emulsion can affect the overall oxidative stability, and possibly also the physical stability, of the emulsion system. Indeed, there have been reports that peptides could suppress the antioxidative action of phenolic compounds.17,18 In our previous study, we have reported that radicalscavenging pea protein hydrolysates (PPHs) prepared with Flavourzyme (Fla-PPH) and Protamex (Pro-PPH) exhibit remarkable synergistic antioxidative effect with licorice extract (LE) (an ethanol extract from the herb Glycyrrhiza glabra) in a liposomal model system.15 However, it is not known whether there exists a similar cooperative inhibitory effect of PPH with LE on lipid oxidation in an O/W emulsion and, if so, how they would partition and interact at the oil−water interface. Therefore, the present study was conducted to investigate the influence of LE on the antioxidant activity of Fla-PPH and ProReceived: Revised: Accepted: Published: 8204
April 4, 2014 July 23, 2014 July 24, 2014 July 24, 2014 dx.doi.org/10.1021/jf5016126 | J. Agric. Food Chem. 2014, 62, 8204−8213
Journal of Agricultural and Food Chemistry
Article
procedure using sodium thiosulfate (Na2S2O3) titration. TBARS were tested according to the method of Mei et al.23 and calculated from the standard curve prepared by 1,1,3,3-tetraethoxypropane. Synergism of PPHs and LE in inhibiting peroxides or TBARS formation was expressed and calculated as
PPH in a soybean O/W emulsion. To elucidate the reaction mechanism, the adsorption of PPH and LE at the emulsion interface and the interfacial properties were examined.
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MATERIALS AND METHODS
Materials. Pea protein isolate (PPI) was extracted from pulverized yellow peas (Pisum sativum L.) using the isoelectric precipitation method that involved initial extraction at pH 8.0 followed by precipitation at pH 4.5.19 After neutralization (pH 7.0), the PPI suspension was lyophilized to yield a dry powder with 92% protein. Flavourzyme and Protamex were donated by Novozymes North America Inc. (Franklinton, NC, USA). Soybean oil was purchased from a local supermarket (Lexington, KY, USA). Licresse, a licorice (G. glabra) root ethanol extract (LE) powder with a 23% total phenolic content and
Synergy of Licorice Extract and Pea Protein Hydrolysate for Oxidative Stability of Soybean Oil-in-Water Emulsions Xin Zhang,† Youling L. Xiong,*,‡ Jie Chen,† and Lirong Zhou§ †
State Key Laboratory of Food Science and Technology and School of Food Science and Technology, Jiangnan University, Wuxi, Jiangsu 214122, China ‡ Department of Animal and Food Sciences, University of Kentucky, Lexington, Kentucky 40546, United States § ICL Performance Products LP, St. Louis, Missouri 63141, United States ABSTRACT: Previously developed radical-scavenging pea protein hydrolysates (PPHs) prepared with Flavourzyme (Fla-PPH) and Protamex (Pro-PPH) were used as cosurfactants with Tween 20 to produce soybean oil-in-water (O/W) emulsions, and the suppression of lipid oxidation was investigated. Both PPHs significantly retarded oxidation (P < 0.05) of the emulsions when stored at 37 °C for 14 days. Electron microscopy revealed an interfacial peptidyl membrane around oil droplets, which afforded steric restrictions to oxidation initiators. When licorice extract (LE) was also used in emulsion preparation, a remarkable synergistic oxidation inhibition was observed with both PPHs. LE adsorbed onto oil droplets either directly or through associating with PPH to produce a thick and compact interfacial membrane enabling the defense against oxygen species. Liquiritin apioside, neolicuroside, glabrene, and 18β-glycyrrhetic acid were the predominant phenolic derivatives partitioning at the interface and most likely the major contributors to the notable synergistic antioxidant activity when coupled with PPHs. KEYWORDS: antioxidant, pea protein hydrolysate, licorice extract, emulsion, oxidative stability, synergism
■
INTRODUCTION A wide array of natural and processed foods exist as emulsions, such as milk, cream, mayonnaise, and salad dressings. Due to the large contact surface between oxidizable fatty acids and water-soluble oxidants (oxygen and transition metal ions), emulsified oil is much more susceptible to oxidation than bulk oil.1 Peptides and protein hydrolysates as potential alternatives to traditional antioxidants have been widely reported to be inhibitory of lipid oxidation in biphasic (O/W) systems through scavenging free radicals, sequestrating pro-oxidative metal ions, or forming physical (shielding) barrier around oil droplets.2−4 Unlike small-molecule antioxidants, protein hydrolysates may also act as stabilizing agents to improve the physical stability of emulsion systems.5,6 Phenolic compounds extracted from plants and spices are also recognized for their strong antioxidant activity in food systems due to radical-scavenging activity and chelation of transition metal ions.7,8 Many polyphenols and their plant extracts are effective against both lipid peroxidation and protein oxidative changes in model systems and in complex food matrices, such as processed meats.9,10 The effectiveness of an antioxidant in retarding lipid oxidation in food emulsions depends not only on its chemical reactivity with oxidation initiators and propagators but also on its location, which is dictated by its polarity and affinity for other constituents.11,12 Due to the size reduction and increased charges, peptides are distributed preferentially in the aqueous phase and can partition at the oil−water interface, thereby preventing the initiation of lipid oxidation induced by watersoluble oxidants. On the other hand, as polyhydroxyl phenol derivatives with both polar and nonpolar moieties, phenolic compounds have the propensity to concentrate at the oil−water interface and are dispersible in the lipid phase (where © 2014 American Chemical Society
hydroperoxides and peroxide radicals are generated) to curtail the propagation of lipid radical chain reactions. Because of their complementary processes in inhibiting lipid oxidation, peptides (protein) and phenolic compounds when used in combination have been found to function synergistically in several biphasic systems, such as O/W emulsions7,13,14 and a liposome suspension.15 However, it should be noted that similar to Tween, Span, and monoglycerides, phenolic molecules as surface active compounds might influence the adsorption of peptides at the interface either indirectly by competing for the oil droplets or directly by binding to peptides through hydrophobic interaction and hydrogen bonds.16 Therefore, the existence of phenolics in a peptide-stabilized emulsion can affect the overall oxidative stability, and possibly also the physical stability, of the emulsion system. Indeed, there have been reports that peptides could suppress the antioxidative action of phenolic compounds.17,18 In our previous study, we have reported that radicalscavenging pea protein hydrolysates (PPHs) prepared with Flavourzyme (Fla-PPH) and Protamex (Pro-PPH) exhibit remarkable synergistic antioxidative effect with licorice extract (LE) (an ethanol extract from the herb Glycyrrhiza glabra) in a liposomal model system.15 However, it is not known whether there exists a similar cooperative inhibitory effect of PPH with LE on lipid oxidation in an O/W emulsion and, if so, how they would partition and interact at the oil−water interface. Therefore, the present study was conducted to investigate the influence of LE on the antioxidant activity of Fla-PPH and ProReceived: Revised: Accepted: Published: 8204
April 4, 2014 July 23, 2014 July 24, 2014 July 24, 2014 dx.doi.org/10.1021/jf5016126 | J. Agric. Food Chem. 2014, 62, 8204−8213
Journal of Agricultural and Food Chemistry
Article
procedure using sodium thiosulfate (Na2S2O3) titration. TBARS were tested according to the method of Mei et al.23 and calculated from the standard curve prepared by 1,1,3,3-tetraethoxypropane. Synergism of PPHs and LE in inhibiting peroxides or TBARS formation was expressed and calculated as
PPH in a soybean O/W emulsion. To elucidate the reaction mechanism, the adsorption of PPH and LE at the emulsion interface and the interfacial properties were examined.
■
MATERIALS AND METHODS
Materials. Pea protein isolate (PPI) was extracted from pulverized yellow peas (Pisum sativum L.) using the isoelectric precipitation method that involved initial extraction at pH 8.0 followed by precipitation at pH 4.5.19 After neutralization (pH 7.0), the PPI suspension was lyophilized to yield a dry powder with 92% protein. Flavourzyme and Protamex were donated by Novozymes North America Inc. (Franklinton, NC, USA). Soybean oil was purchased from a local supermarket (Lexington, KY, USA). Licresse, a licorice (G. glabra) root ethanol extract (LE) powder with a 23% total phenolic content and
