E2 protein nanocage as a pickering emulsifier and its applications

• Main Author: Sarker, Mridul
• Other Authors: School of Chemical and Biomedical Engineering
• Format: Theses and Dissertations
• Language: English
• Published: 2018
• Subjects: DRNTU::Engineering::Bioengineering
• Online Access: http://hdl.handle.net/10356/73892
• Institution: Nanyang Technological University

Particle-stabilized emulsion or Pickering emulsion has drawn attention in recent years because of its several attractive features. Pickering emulsion offers higher stability in different conditions and it has found applications in a variety of industries including food, pharmaceuticals, and consumer products. The selection of particles or Pickering emulsifier is crucial to engineering the characteristic features of Pickering emulsion. Inorganic, organic as well as hybrid nanoparticles have been used in stabilizing Pickering emulsions for different applications with some identified limitations. Inorganic particle is less preferred for the development of food and pharmaceutical products due to its limited biocompatibility. Organic particles have low stability in different conditions presenting a research gap and a quest for new organic materials to overcome these issues. In this work, a protein-based material, E2 protein nanocage, has been introduced as a new Pickering emulsifier. E2 is the first non-viral protein nanocage to be used in stabilizing Pickering emulsion. E2 exhibits excellent surface activity because of the amphiphilic nature of its external surface. E2 adsorbs at the liquid-liquid interface and provides kinetic stability to the emulsion. The E2-stabilized emulsion exhibits pH-switchability for at least 5 cycles and shows stability in a wide range of pH (neutral to basic), ionic strength (as high as 250 mM NaCl) and storage temperature (up to 50°C). The increase in oil fraction in the emulsion composition results in the formation of gel-like structure. The rheological analysis of Pickering emulsion and gel-like structure confirmed the higher mechanical strength, long-term stability and elasticity of the gel-like structure compared to the emulsion. The structural integrity of E2 at the oil/water interface has been studied experimentally and theoretically. The adsorption kinetics of E2 on model hydrophobic surface confirms the formation of 26 nm in a thick monolayer of protein nanocage. In-depth study of the molecular structure reveals an intact tertiary structure of E2 at the interface while the secondary structure results in an increase in alpha-helicity. The trends of denaturation of E2 protein in different denaturation conditions confirm the trend of structural change of E2 at the interface. Furthermore, the theoretical analysis suggests that hydrophobic force dominates over surface force indicating the structural integrity of protein cage at the interface. To explore a potential application in formulating functional food products, of E2-stabilized Pickering emulsion has been proposed as a platform to deliver active molecules in the gastrointestinal tract with two-step release mechanism. Micronutrients such as iron or calcium can be loaded in the E2 while lipophilic micronutrients in the oil phase of the emulsion. The digestion profile of E2 and E2-stabilized emulsion has been investigated in-vitro using gastric simulated fluid followed by the analysis of the micronutrients bioaccessibility in the first step in stomach phase. In summary, E2 has been shown to stabilize emulsion with pH-responsive behavior. Examination of its structure suggests that it maintains a cage-like structure at the oil/water interface. The non-viral origin combined with the loading of active molecules in both the E2 interior and the oil phase makes E2-stabilized Pickering emulsion attractive as nanocarriers with two-step release mechanism for functional food formulation.