Unravelling the structure and dynamics of lipid nanoparticles for biomolecule delivery

Encapsulation in self-assembled lipid structures has been shown to increase the stability of large biomolecules, including proteins and nucleic acids (NAs), which has many potential applications in the food and pharmaceutical industries. In order to fully optimise the encapsulation system for a certain application, however, it is important to understand the effect of the encapsulated biomolecule on the lipid structure and vice versa.
In the first part of my thesis, the encapsulation of different proteins in a lipid sponge phase (L3) system composed of food grade lipids was studied. Comparison between the effect on the L3 phase of encapsulated beta-galactosidase (238 kDa), aspartic protease (34 kDa), myoglobin (17.6 kDa) and phytoglobin BvPgb 1.2 (38.4 kDa) demonstrated the importance of specific lipid-protein interactions on the L3 structure both in bulk and dispersions, as well as the stability of the encapsulated proteins. Systematic variation of the buffer and pH used to prepare and disperse the bulk L3 phase highlighted pH and specific buffer effects on the L3 structure.
In the second part of my thesis, the structure of lipid nanoparticles (LNPs) for NA delivery was investigated using scattering methods and cryogenic transmission electron microscopy (cryoTEM). mRNA adsorption with pH and proportion of the cationic ionisable lipid (CIL) DLin-MC3-DMA was first studied in a model system for two model mRNAs and human erythropoietin mRNA. Effect of nucleic acid type and concentration was characterised in LNPs formulated with a benchmark LNP composition and four different NAs (polyadenylic acid, polyuridylic acid, double stranded and single stranded DNA). Using a combination of small angle x-ray and neutron scattering (SAX/NS), dynamic light scattering (DLS) and cryoTEM, the colloidal stability, LNP morphology, lipid composition of the LNP core-shell structure and structure of the core were determined.