Pea Protein Based Nanoemulsions for Delivery of Vitamin D: Fabrication, Stability and In vitro Study using Caco-2 Cells

• Author / Creator

  Walia, Niharika

• In North America, a significant population is vitamin D deficient due to insufficient sunlight exposure in winters. Increasing research has shown that vitamin D, apart from its skeletal functions, also has potential to lower the risk of chronic diseases such as autoimmune disorders, diabetes mellitus, cardiovascular diseases, and cancers. However, vitamin D exhibits low bioavailability due to its poor solubility in human gut. This research aims to develop pea protein stabilized nanoemulsions as vitamin D delivery systems for improved vitamin absorption.
  In the first study, nanoemulsions stabilized by pea protein were developed using a high-energy approach by high-pressure homogenization treatment. The nanoemulsions exhibited controllable sizes ranging from 170 to 350 nm, good stability with zeta-potential value of around -25mV, and high vitamin D encapsulation efficiency of 94-96%. Using Caco-2 cell model, the nanoemulsions were found relatively safe after 6 hours of incubation with cell viability >80%. Cellular uptake efficiency of small sized nanoemulsions (233 nm) was found to be ~ 2.5 folds higher (p < 0.05) than large sized nanoemulsions (350 nm). Interestingly, protein based nanoemulsions exhibited significantly higher cellular uptake than emulsions prepared using a combination of protein and lecithin. The transport efficiency of vitamin D across Caco-2 cells for small sized nanoemulsions (233 nm) was ~ 5.3 times greater than free vitamin D suspension.
  In the second study, a low-energy approach was used to prepare a complex nanoemulsion formed by a combination of Tween 80 and pea protein by spontaneous emulsification method, which does not rely on high-energy equipment and thus, the processing can be more convenient, cost effective and environmentally sustainable. Tween 80 (3% w/v) was used as a small molecular surfactant to prepare nanoemulsion droplets of 134.8 nm, followed by addition of pea protein (3% w/v), resulting in the formation of complex nanoemulsions with particle size of 207.7 nm. Infrared spectroscopy was used to investigate the effect of Tween 80 on protein conformation, and Tween 80 showed partial denaturation of pea protein molecules. A “clusters on string” structure was proposed for the complex nanoemulsions, which was supported by transmission electron microscopy and infrared spectroscopy, revealing similar particle size as measured by light scattering, and featuring Tween 80 stabilized micelles clustered together on pea protein polymeric chains. Encapsulation efficiencies of vitamin D were as high as 89.3% and 90.1% for nanoemulsions stabilized by Tween80-pea protein and Tween 80 alone. Such complex nanoemulsions were found to remain physically stable during storage for two weeks. The in vitro assays using Caco-2 cells indicated that both nanoemulsions stabilized by Tween 80 alone and the complex nanoemulsion could efficiently improve vitamin D uptake by ≥ 2.5 folds (p < 0.05) as compared to free vitamin D suspension. Interestingly, the transport efficiency of vitamin D across Caco-2 cells for complex nanoemulsions was 5.6 folds higher than free vitamin D suspension and 2.3 folds higher than nanoemulsions stabilized by Tween alone.
  As a result, these pea protein stabilized nanoemulsions demonstrated efficient encapsulation, cellular uptake and transport of vitamin D using Caco-2 cell model to simulate absorption through intestinal epithelium. Such protein-based nanoemulsions may allow more efficient vitamin D delivery due to its small particle size to improve the status of vitamin deficiencies in aged population and in areas with less sun exposure. Moreover, these nanoemulsions can be developed by both high-energy approach or low-energy approach with the help of natural large molecular surfactants (pea protein) or a combination of pea protein and a low amount of small molecular surfactant (Tween 80), respectively. The former has a “surfactant free” status, and thus appeals more to food industries and gives brand a cleaner label; while the latter is more interesting due to its cost effective and environmentally sustainable way of nanoemulsions production. Moreover, pea protein is an emerging protein source of plant origin and not regarded as a major allergen, and thus the developed nanoemulsion delivery system of vitamin D has potential applications to create novel non-dairy functional foods and beverages suitable for different populations such as lactose intolerant, vegans or consumers with little preference for dairy.

• Subjects / Keywords

  • Delivery system
  • Nanoemulsion
  • Vitamin D
  • Encapsulation
  • Caco-2 cells
  • Functional food
  • Tween 80
  • Plant protein
  • Pea protein

• Graduation date

  Fall 2019

• Type of Item

  Thesis

• Degree

  Master of Science

• DOI

  https://doi.org/10.7939/r3-xp81-ys04

• License

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