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Cite this: Food Funct., 2014, 5, 65
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Responsiveness of emulsions stabilized by lactoferrin nano-particles to simulated intestinal conditions
Published on 28 October 2013. Downloaded on 27/10/2014 19:18:05.
Dafna Meshulamab and Uri Lesmes*ab There is an upsurge of interest in the use of nano-particles to fabricate emulsions and modulate their functionality, with particular emphasis on modulating emulsion digestive fate. Food grade nano-particles formed through controlled processing and electrostatic biopolymer interactions are yet to be systematically studied for their ability to stabilize emulsions and modulate emulsion digestibility. This study focused on the responsiveness of emulsions stabilized by lactoferrin (LF) nano-particles (NPs) and dietary fibers to key digestive parameters. Compared to native LF, LF-NPs comprised emulsion exhibited elevated creaming rates as evident from accelerated stability tests performed by analytical centrifugation. The electrostatic deposition of alginate or carrageenan onto the LF-NPs significantly improved the stability of the corresponding emulsions. Further, the use of various nano-particles showed to have both beneficial and deleterious effects on emulsion responsiveness to pH (2.0 < pH < 10.0), CaCl2 (0–40 mM) and bile (0–25 mg mL1). Simulated pH-stat lipolysis experiments show that the use of LF or LF-NPs had no marked effect on lipolysis. Intriguingly, the use of LF-NPs and alginate reduced emulsion lipolysis by 14% while the use of LF-NPs and carrageenan increased lipolysis by 10%. Microscopy images as well as Received 2nd September 2013 Accepted 27th October 2013
droplet characterization in terms of size and charge indicate that the altered emulsion responsiveness may be due to physical differences in emulsion properties (e.g. droplet size) and overall organization
DOI: 10.1039/c3fo60380f
during digestion (e.g. aggregation vs. coalescence). Overall, this study’s insights could prospectively be
www.rsc.org/foodfunction
used to harness protein nano-particles to tweak emulsion behavior during digestion.
1. Introduction Functional foods are a vivid eld of research seeking to nutritionally intervene, alleviate or prevent modern morbidity, such as obesity, coronary heart disease and diabetes. In recent years, there has been an upsurge of interest in rational design and product engineering to incorporate more bioactive ingredients into our diet, to maximize the advantages of food processing and enhance food’s contribution to human health exceeding its nutritional value.1 Emulsions are extremely common in foods and have been enthusiastically studied for their potential to serve as efficient and cost effective delivery systems for lipophilic bioactive components.1–5 However, emulsions’ inherent thermodynamic instability renders the use of emulsiers and/or stabilizers a practical necessity in the manufacturing of kinetically stable emulsions.5 Thus, the modication of emulsiers and of droplet interfacial properties has been documented to
a
Laboratory of Chemistry of Foods and Bioactives, Department of Biotechnology and Food Engineering, Technion – Israel Institute of Technology, Haifa 32000, Israel. E-mail: [email protected]; Fax: +972-4-8293399; Tel: +972-77-8871869 Russel Berrie Nanotechnology Institute, Technion – Israel Institute of Technology, Haifa 32000, Israel
b
This journal is © The Royal Society of Chemistry 2014
affect emulsions’ susceptibility to various stresses and even to manipulate their digestive fate.3,5–8 The ever existing need to overcome the limitations of current emulsion formulations maintains a constant need for innovations in the eld. Recently, the use of solid colloids to form and stabilize emulsions, also termed Pickering emulsions, has been increasingly described in the literature.9,10 These unique colloidal systems offer the possibility to fabricate emulsions with distinctly improved physical and chemical stability, even from foodcomplaint colloids, such as polysaccharide nano-particles (NPs).9,11–13 Such emulsions are uniquely stabilized by colloids practically irreversibly adsorbed onto the droplet interface providing marked steric hindrance.14 The effectiveness of colloids to generate such unique physical barriers depends on various parameters such as oil type, particle size, inter-particle interactions, and particle wettability.9,15 Various food compliant hydrocolloids have recently been described in the literature, including starch granules, chitin nano-crystals, cellulose nano-crystals, solid lipid nanoparticles and brils.9–13,16–20 Protein–polysaccharide complex NPs formed through electrostatic biopolymer interactions have also been recently described as potential Pickering emulsion stabilizers.21 The formation and stability of such structured food NPs have even been recently reviewed.22 Moreover, the incorporation of
Food Funct., 2014, 5, 65–73 | 65
View Article Online
Published on 28 October 2013. Downloaded on 27/10/2014 19:18:05.
Food & Function
proteins into emulsion systems is thought to potentially offer additional health benets, as some alimentary proteins have been demonstrated to have specic bioactivities, such as antimicrobial, antioxidant, antihypertensive or immune-modulating activities.23,24 In addition, certain polysaccharides have also been shown to possess bioactivity. For example, indigestible carbohydrates classied as dietary bers are associated with many potential health benets, including modulation of colon ora, prevention of some cancer types and are even thought of as therapeutic targets tackling obesity and metabolic syndromes.25,26 Thus, various bioactive proteins and carbohydrates in emulsions can be used to form multifunctional emulsions in which the lipid droplet may deliver lipophilic bioactives and the formulation stabilizers may provide additional bioactive components to the formulation. Further, the integration of proteins and polysaccharides into emulsion formulations can be efficient tools in modulating the droplet interfacial composition and properties and consequently modulating emulsion digestibility in the human gastrointestinal tract (GIT).27–29 Still, little is known on the role of protein-based NPs in stabilizing emulsions and modulating emulsion responsiveness to the conditions of the GIT which govern the emulsion digestive fate. Recent studies have demonstrated the conditions and properties of different lactoferrin nano-particles formed via thermal processing and pH treatments, based on the high isoelectric point of lactoferrin (pI > 8.0).30–32 Such LF-based NPs have also been shown to exhibit altered susceptibility to gastric proteolysis.29 Most recently, we have demonstrated that emulsions stabilized by certain LF-based NPs may exhibit altered responsiveness to articial saliva and in vitro gastric digestion.21 Overall, these studies have shown that certain polysaccharides can interfere with gastrointestinal proteolysis and at the same time be useful as emulsiers yielding emulsions with varying functionalities and susceptibility to oral and gastric conditions. Thus, the present study was dedicated to expand our understanding on the responsiveness of emulsions stabilized by lactoferrin–dietary ber NPs to key physiological parameters of the small intestine.
2.
Materials and methods
2.1. Materials Food grade bovine lactoferrin (Vivinal lactoferrin FD, 95.6% protein,
PAPER
Cite this: Food Funct., 2014, 5, 65
View Journal | View Issue
Responsiveness of emulsions stabilized by lactoferrin nano-particles to simulated intestinal conditions
Published on 28 October 2013. Downloaded on 27/10/2014 19:18:05.
Dafna Meshulamab and Uri Lesmes*ab There is an upsurge of interest in the use of nano-particles to fabricate emulsions and modulate their functionality, with particular emphasis on modulating emulsion digestive fate. Food grade nano-particles formed through controlled processing and electrostatic biopolymer interactions are yet to be systematically studied for their ability to stabilize emulsions and modulate emulsion digestibility. This study focused on the responsiveness of emulsions stabilized by lactoferrin (LF) nano-particles (NPs) and dietary fibers to key digestive parameters. Compared to native LF, LF-NPs comprised emulsion exhibited elevated creaming rates as evident from accelerated stability tests performed by analytical centrifugation. The electrostatic deposition of alginate or carrageenan onto the LF-NPs significantly improved the stability of the corresponding emulsions. Further, the use of various nano-particles showed to have both beneficial and deleterious effects on emulsion responsiveness to pH (2.0 < pH < 10.0), CaCl2 (0–40 mM) and bile (0–25 mg mL1). Simulated pH-stat lipolysis experiments show that the use of LF or LF-NPs had no marked effect on lipolysis. Intriguingly, the use of LF-NPs and alginate reduced emulsion lipolysis by 14% while the use of LF-NPs and carrageenan increased lipolysis by 10%. Microscopy images as well as Received 2nd September 2013 Accepted 27th October 2013
droplet characterization in terms of size and charge indicate that the altered emulsion responsiveness may be due to physical differences in emulsion properties (e.g. droplet size) and overall organization
DOI: 10.1039/c3fo60380f
during digestion (e.g. aggregation vs. coalescence). Overall, this study’s insights could prospectively be
www.rsc.org/foodfunction
used to harness protein nano-particles to tweak emulsion behavior during digestion.
1. Introduction Functional foods are a vivid eld of research seeking to nutritionally intervene, alleviate or prevent modern morbidity, such as obesity, coronary heart disease and diabetes. In recent years, there has been an upsurge of interest in rational design and product engineering to incorporate more bioactive ingredients into our diet, to maximize the advantages of food processing and enhance food’s contribution to human health exceeding its nutritional value.1 Emulsions are extremely common in foods and have been enthusiastically studied for their potential to serve as efficient and cost effective delivery systems for lipophilic bioactive components.1–5 However, emulsions’ inherent thermodynamic instability renders the use of emulsiers and/or stabilizers a practical necessity in the manufacturing of kinetically stable emulsions.5 Thus, the modication of emulsiers and of droplet interfacial properties has been documented to
a
Laboratory of Chemistry of Foods and Bioactives, Department of Biotechnology and Food Engineering, Technion – Israel Institute of Technology, Haifa 32000, Israel. E-mail: [email protected]; Fax: +972-4-8293399; Tel: +972-77-8871869 Russel Berrie Nanotechnology Institute, Technion – Israel Institute of Technology, Haifa 32000, Israel
b
This journal is © The Royal Society of Chemistry 2014
affect emulsions’ susceptibility to various stresses and even to manipulate their digestive fate.3,5–8 The ever existing need to overcome the limitations of current emulsion formulations maintains a constant need for innovations in the eld. Recently, the use of solid colloids to form and stabilize emulsions, also termed Pickering emulsions, has been increasingly described in the literature.9,10 These unique colloidal systems offer the possibility to fabricate emulsions with distinctly improved physical and chemical stability, even from foodcomplaint colloids, such as polysaccharide nano-particles (NPs).9,11–13 Such emulsions are uniquely stabilized by colloids practically irreversibly adsorbed onto the droplet interface providing marked steric hindrance.14 The effectiveness of colloids to generate such unique physical barriers depends on various parameters such as oil type, particle size, inter-particle interactions, and particle wettability.9,15 Various food compliant hydrocolloids have recently been described in the literature, including starch granules, chitin nano-crystals, cellulose nano-crystals, solid lipid nanoparticles and brils.9–13,16–20 Protein–polysaccharide complex NPs formed through electrostatic biopolymer interactions have also been recently described as potential Pickering emulsion stabilizers.21 The formation and stability of such structured food NPs have even been recently reviewed.22 Moreover, the incorporation of
Food Funct., 2014, 5, 65–73 | 65
View Article Online
Published on 28 October 2013. Downloaded on 27/10/2014 19:18:05.
Food & Function
proteins into emulsion systems is thought to potentially offer additional health benets, as some alimentary proteins have been demonstrated to have specic bioactivities, such as antimicrobial, antioxidant, antihypertensive or immune-modulating activities.23,24 In addition, certain polysaccharides have also been shown to possess bioactivity. For example, indigestible carbohydrates classied as dietary bers are associated with many potential health benets, including modulation of colon ora, prevention of some cancer types and are even thought of as therapeutic targets tackling obesity and metabolic syndromes.25,26 Thus, various bioactive proteins and carbohydrates in emulsions can be used to form multifunctional emulsions in which the lipid droplet may deliver lipophilic bioactives and the formulation stabilizers may provide additional bioactive components to the formulation. Further, the integration of proteins and polysaccharides into emulsion formulations can be efficient tools in modulating the droplet interfacial composition and properties and consequently modulating emulsion digestibility in the human gastrointestinal tract (GIT).27–29 Still, little is known on the role of protein-based NPs in stabilizing emulsions and modulating emulsion responsiveness to the conditions of the GIT which govern the emulsion digestive fate. Recent studies have demonstrated the conditions and properties of different lactoferrin nano-particles formed via thermal processing and pH treatments, based on the high isoelectric point of lactoferrin (pI > 8.0).30–32 Such LF-based NPs have also been shown to exhibit altered susceptibility to gastric proteolysis.29 Most recently, we have demonstrated that emulsions stabilized by certain LF-based NPs may exhibit altered responsiveness to articial saliva and in vitro gastric digestion.21 Overall, these studies have shown that certain polysaccharides can interfere with gastrointestinal proteolysis and at the same time be useful as emulsiers yielding emulsions with varying functionalities and susceptibility to oral and gastric conditions. Thus, the present study was dedicated to expand our understanding on the responsiveness of emulsions stabilized by lactoferrin–dietary ber NPs to key physiological parameters of the small intestine.
2.
Materials and methods
2.1. Materials Food grade bovine lactoferrin (Vivinal lactoferrin FD, 95.6% protein,
