Purpose: Attempts to develop polymer-lipid hybrid nanoparticles have shown that these nanoparticles have the beneficial features of both lipid nanoparticles and polymer nanoparticles. More recently, protein-based nanocarriers have emerged in the pharmaceutical market due to their excellent biodegradability and biocompatibility. Zein is a hydrophobic and amphiphilic protein obtained from maize. Zein is used in the fields of food science and biomedicine due to its FDA generally recognized as safe (GRAS) status and ease of preparation into spherical colloidal particles. To explore the features of protein-lipid hybrid nanoparticles as drug carriers, in the present study we sought to develop such nanoparticles with zein as the protein component and phosphatidylcholine (PC) as the lipid component.
Methods: Zein-PC hybrid nanoparticles (Z/PC-NP) were prepared by freeze-drying method. Incorporated zein concentration was determined by measuring the protein absorbance at 278 nm using spectrophotometer. Differential scanning calorimetry (DSC) was used to investigate the effect of zein incorporation on phase transition behavior of PC. Size and morphology of various nanoparticles (Z/PC-NP, zein nanoparticles (Z-NP) and liposomes (PC-NP)) were determined by dynamic light scattering method and negative staining and cryogenic transmission electron microscopy. Drug loaded concentration in various nanoparticles was determined by HPLC analysis (paclitaxel, docetaxel, celecoxib) and spectrophotometry (curcumin). Storage stability (4oC) and serum condition stability (37oC) of nanoparticles were evaluated by the changes in the mean size of nanoparticles. In addition, lyophilization property of nanoparticles was assessed by the changes in the mean particle size and retained drug content. Docetaxel release profile of various nanoparticles reconstituted after lyophilization was assessed in physiologically relevant condition.
Results: The incorporated zein amount of nanoparticles increased as the initial zein was increased from 2.5 mg to 10 mg while the zein incorporation efficiency decreased. Z/PC-NPs, containing DOPC:EDOPC as a PC revealed higher zein incorporation efficiency than DMPC or DOPC alone nanoparticles. DSC data indicated that zein was incorporated within phosphatidylcholine bilayer. Z/PC-NP, containing DOPC:EDOPC as a PC, were spherical particles with <200 nm and exhibited highly positive surface charge. Cryo-TEM image showed that the shell composed of the mixture of zein and PC surrounded the spherical zein core structure. Among Z/PC-NP, Z-NP and PC-NP, Z/PC-NP exhibited the highest drug-loading efficacy in cases of all four drugs and higher stability in serum condition than PC-NP. In addition, cholesterol incorporation greatly decreased the retained paclitaxel content in PC-NP after lyophilization of nanoparticles but not that in Z/PC-NP. The time-dependent release of docetaxel was faster from Z/PC/CHOL NP compared to PC/CHOL-NP but much slower than free docetaxel.
Conclusion: The formed Z/PC-NP appeared to be spherical with nanometer dimensions, and each composed of a zein core surrounded by a zein/PC hybrid shell. The improved features of Z/PC-NP compared to the non-hybrid counterparts are expected to provide an advanced hydrophobic drug carrier with promising potential.