Characterization of Nanohybridosomes from Lipids and Spruce Homogenate Containing Extracellular Vesicles

Lipid nanovesicles associated with bioactive phytochemicals from spruce needle homogenate (here called nano-sized hybridosomes or nanohybridosomes, NSHs) were considered.

We formed NSHs by mixing appropriate amounts of lecithin, glycerol and supernatant of isolation of extracellular vesicles from spruce needle homogenate. We visualized NSHs by light microscopy and cryogenic transmission electron microscopy and assessed them by flow cytometry, dynamic light scattering, ultraviolet-visual spectroscopy, interferometric light microscopy and liquid chromatography-mass spectrometry.

We found that the particles consisted of a bilayer membrane and a fluid-like interior. Flow cytometry and interferometric light microscopy measurements showed that the majority of the particles were nano-sized. Dynamic light scattering and interferometric light microscopy measurements agreed well on the average hydrodynamic radius of the particles R_h (between 140 and 180 nm), while the concentrations of the particles were in the range between 10¹³ and 10¹⁴/mL indicating that NSHs present a considerable (more than 25%) of the sample which is much more than the yield of natural extracellular vesicles (EVs) from spruce needle homogenate (estimated less than 1%). Spruce specific lipids and proteins were found in hybridosomes.

Simple and low-cost preparation method, non-demanding saving process and efficient formation procedure suggest that large-scale production of NSHs from lipids and spruce needle homogenate is feasible.