Potential applications of lipid nanoparticles in edible packaging and nutraceutical delivery
Creators: name: Palaparthi, Annie D'Souza
Publisher: name: No Publisher Supplied
Year: 2016

description: Lipid nanoparticles are emulsion-type nanoscale aqueous dispersions that have been extensively used as delivery systems for drugs and active ingredients of lipophilic nature in pharmaceutical and cosmetic industry through different administration routes. These nanoparticles are made from food-grade ingredients which make them desirable approach to create new fortified and functional foods.    In my work, lipid nanoparticles were investigated for the very first time as a potential nanofiller in biopolymer films in improving moisture barrier properties and further extended their role in creating multifunctional edible films. Further, a new lipid nanoparticle/nanocarrier from cocoa butter was developed and evaluated as an effective delivery system of non-polar bioactives for functional beverage applications. Finally, this nanocarriers were tested as nanofillers in biopolymer films that improve its moisture interactions as an edible film and examine whether these films can serve an "encapsulating matrices" to delivery systems.    Candelilla wax, a highly hydrophobic plant wax was utilized to create a nano dispersion using microemulsion technique. Selection of emulsifiers and proportions of the emulsions were carried out by using effective HLB concept for emulsifiers and ternary phase diagrams respectively. Particle size analysis using dynamic light scattering and imaging of Candelilla wax nanoparticles using atomic force microscopy were examined to understand their storage stability and morphology. Chitosan, an antimicrobial natural polymer was used as a model nanocomposite film to analyze its physical properties and antibacterial nature with wax nanoparticles. The film was preparing film solution by blending polyvinyl alcohol, glycerol and wax nanoparticles to create a thin film which can be used for practical purposes. After conditioning at ambient conditions (relative humidity of 53% and room temperature 25Â°C, moisture barrier properties (surface hydrophilicity, % water uptake, moisture isotherms and water vapor permeability) of the films were studied. Mechanical strength and elasticity of the films were calculated and microstructure of the surface and cross-sections of the film were examined to understand their physical state. Inclusion of fillers might affect the anti-bacterial nature of chitosan and to understand the effect of wax nanoparticles, growth and interaction of gram-negative and gram-positive bacteria on these films were measured to establish the nanocomposite films' anti-microbial activity. Optimization of preparation of hot emulsions of Candelilla wax with Tween 80 (polysorbate 80) gave nanoparticles of smallest average hydrodynamic diameter of ~ 212 nm with good storage stability. On visualizing the nanoparticle with AFM and cross-sectional analysis, it was found that they have a platelet structure with dimensions ~145nm as the horizontal size and ~15nm as its vertical height. Films were prepared with different percentages of nanoparticles keeping chitosan and glycerol ratio constant.  From surface hydrophilicity studies using water droplet method with contact angle analysis, it was noted that hydrophobicity of the film with wax nanoparticles was slightly increased and at a higher concentrations of nanoparticles (&gt; 3% w/w), contact angle with water did not change. From moisture isotherms of films with different concentrations of nanoparticles, adsorption of water was significantly decreased as the movement of water molecules were restricted in nanocomposites. Water uptake capacity of the nanocomposites were drastically lowered as percentage of nanoparticles in the film increased. On increasing nanoparticle content from 1% to 10% w/w, water vapor permeability values decreased by 19% to 55%. Tensile strength and elongation % values of nanocomposites were compared with the control film without nanoparticles, showing that nanoparticles have no plasticizing effect. Effects of wax nanoparticles in the films were further supported by its homogenous structure and chemical interaction analysis by FTIR. Antibacterial nature of chitosan was not affected at low content (&lt; 5% w/w) of nanoparticles. From these observations, it can be concluded that Candelilla wax nanoparticles can serve as potential nanofiller in chitosan films and these nanocomposites can effectively be used as primary packaging material as stable over a wide range of relative humidity and increase shelf-life with its controlled water transfer and anti-microbial nature.  Health-promoting bioactives due to their chemical nature are not effective due to their instability, poor bioaccessibility and bioavailability on digestion. In pharmaceutical industry, lipids based delivery systems have shown to overcome this problem. Lipid based nanoparticles, also called as nanocarriers were developed to deliver beneficial non-polar molecules that promote wellness and alleviate diseases conditions. In my study, emulsion based nanoparticles are prepared from cocoa butter with Coenzyme Q10 (CoQ10) as a test bioactive which has poor bioavailability due to its hydrophobic nature. Selection of lipid for suitable for nanoparticle preparation is dependent on the chemical properties of the bioactive. Solubility of the bioactive is the key factor that governs the encapsulation and stability of a delivery system. Cocoa butter (CB) was chosen due to high solubility of CoQ10 (110 Â± 4.7% w/w) among the soft fats considered for the study and its polymorphic crystal states that enables better loading capacity and stability of nanocarriers. Cocoa butter emulsions were prepared with polysorbate and fatty acid esters based non-ionic emulsifiers using HLB systematic approach and further, ternary phase diagrams were created to narrow down compositions of stable oil-in-water (O/W) emulsions. According to the solubility studies with CoQ10, these stable O/W emulsions were loaded with CoQ10 and particle size analysis was conducted to obtain the most stable formulation with highest loading capacity (2% w/w). These hot emulsions were subjected to homogenization above melting temperatures to create nanoscale emulsion droplets with CoQ10 and quickly cooled to 4Â°C. The process conditions of high pressure homogenization (1000 bar and 20 cycles) were optimized to obtain nanoemulsions of smallest diameter required for maximum saturation solubility and better dissolution velocity of encapsulated CoQ10. To understand the effect of CoQ10 concentration on cocoa butter nanocarriers, three different formulations were prepared 0.5% (low-level), 1.0% (mid-level) and 2.0% (high-level) CoQ10 with 2% w/w emulsifier (1.74% Span60 and 0.36% Tween60), 10% cocoa butter and remaining DI water. Storage stabilities of three formulations show that they are stable with no significant difference in particle size over 7 weeks.  Due to high solubility of CoQ10 in CB, 85-90% entrapment efficiency was achieved in nanocarriers.  Formulations did not destabilize after sterilization which is a crucial HACCP step for beverage applications. Crystallization state of components and structural information of these nanocarriers can throw light on its stability and behavior on consumption. Most of cocoa butter exists a metastable Î± polymorph form in an empty nanocarriers form and in presence of CoQ10, different polymorphs existed. CoQ10 remains as a super-cool melt during recrystallization process as there were no phase transition peak from liquid to solid crystal state. Structural data about nanoparticles can be elucidated from scattering intensity profiles generated by synchrotron small-angle X-ray scattering (SAXS) method. Using Porod's law, structural information extracted from scattering intensity profile shows that the interface thickness is affected by loading % of CoQ10 at intermediate q range. Empty and low load nanocarriers have interface thickness of 4.5 nm and in high load nanocarriers, interface thickness increases by only 9%. This shows that interfaces of nanocarriers is not affected by encapsulate.   Performance of this novel CoQ10-CB nanocarriers by evaluating the bioaccessibility (a measure for amount of solubilized CoQ10 that is available for absorption) and relative bioavailability (a measure for amount of CoQ10 that reaches systemic circulation over time of observation) with in vitro and in vivo models. In vitro lipolysis with pancreatic lipase in fed and fast state buffers were conducted and compared on 0.5%, 1.0% and 2.0% CoQ10 in both mixture and nanoparticle form without changing material composition. Extent of lipolysis is a parameter studied to understand lipase activity on different types of formulations and measure amount of fatty acids released during lipolysis. Upon 2-way ANOVA analysis. there was no significant difference in extent of lipolysis on comparing fed and fast state between mixture and nanoparticle form. CB lipolysis in nanoparticle form was significantly greater than that of mixture due to greater surface area and better exposure to enzyme in nanodispersions.  However, it was interesting to find that addition of CoQ10 in CB mixtures and nanocarriers have significantly lowered the lipolysis of CB by 32-42%. Bioaccessibility of CoQ10 significantly improved when incorporated in cocoa butter nanoparticles. Among different formulations with different levels of CoQ10, formulations with lowest amount of CoQ10 (0.5% CoQ10-CBLN) had highest bioaccessibility suggesting that there is a limiting concentration of CoQ10 in mixed micelles of hydrolysis products of nanocarriers and is controlled by lipase activity on the delivery system...