Non-lamellar lipid aqueous phases, such as reverse cubic or hexagonal phases, can be used to entrap smaller biomolecules. The curvature of these lipid phases and hence the size of the aqueous cavities depends on the composition, water content and temperature. The challenge is to encapsulate proteins, such as large enzymes due to the limited size of these cavities. Here, we will present a lipid system, based on mixtures of acylglycerides and acyldiglycerides, which are able to form highly swollen sponge phases (L3), with aqueous pores up to 13 nm of diameter. The structure and composition of the particles were revealed by using small angle neutron scattering (SANS), light scattering, cryo-TEM, size exclusion chromatography and Raman spectroscopy. The results for the sponge phases are compared with data for lamellar and reverse bicontinuous cubic phase in the same lipid system. Large similarities in lipid chain confirmation and head group interactions are observed as all three structures are formed by lipid bilayers, albeit of different curvature. The L3 structure in excess aqueous solution form sponge-like nanoparticles (L3 NPs). We investigate encapsulation of two key types of enzymes of different sizes, used in food processing, namely Aspartic protease (34 KDa) and Beta-galactosidase (460 KDa). They are today delivered into the process as solutions with a considerable amount of preservatives and still with limited shelf-life and limited control of the enzyme activity. The SANS results reveal differences in the L3 NPs with and without enzyme that can be interpreted as inclusion of the protein in the liquid crystalline phase. These findings are verified by size exclusion chromatography and the enzymatic activity of the encapsulated enzyme, which surpasses the storage stability of pure enzymes in solution. To reveal the nature of the interaction between the enzymes and the lipid matrix, we further studied the adsorption of both proteins on the lipid layers formed by the L3 NPs. These data reveal partial penetration of the enzymes in the lipid bilayers. The lipid self-assembly structure can also be changed in desired way by adding a particular type of lipolytic enzymes. Here the (oil) triglyceride aqueous interface is important also for other processes like emulsification. Our results using ellipsometry and x-ray scattering show that uptake of water in the triolein phase can occur and this can be affected by the presence of lipolytic enzymes.

Lipid self-assembly in bulk and at interfaces: Non-lamellar phases and biomolecular interactions

2019 Spring National Meeting

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