Formulation, characterization and optimization of liposomes containing eicosapentaenoic and docosahexaenoic acids; a methodology approach

ABSTRACT

Omega-3 fatty acids [FAs] have been shown to prevent cardiovascular disease. The most commonly used omega-3 fatty acids like eicosapentaenoic acid [EPA] and docosahexaenoic acid [DHA] are highly vulnerable to oxidation and therefore, have short shelf life. Recent advances in nanoliposomes provided a biocompatible system for stabilizing omega-3 FAs. Several methods could be implemented to prepare nanoliposomes. To the best of our knowledge, the performances of these methods in preparation omega-3 FAs have not been examined. Nanoliposomes were prepared by thin film hydration followed by one of the following methods: 1- extrusion, ultrasonic irradiation; 2- bath sonication; 3- probe sonication; or 4- combined probe and bath sonication. The size of liposomes obtained from methods 1 to 4 were 99.7 +/- 3.5, 381.2 +/- 7.8, 90.1 +/- 2.3, and 87.1 +/- 4.10 nm with zeta potential being -42.4 +/- 1,7,-36.3 +/- 1.6, -43.8 +/- 2.4, and 31.6 +/- 1.9 mV, respectively. The encapsulation efficiency [EE] for DHA was 13.2 +/- 1.1%, 26.7 +/- 1.9%, 56.9 +/- 5.2% and 51.8 +/- 3.8% for methods 1 to 4, respectively. The corresponding levels for EPA were 6.5 +/- 1.3%, 18.1 +/- 2.3%, 38.6 +/- 1.8%, and 38 +/- 3.7%, respectively. The EE for DHA and EPA of liposomes for both methods 3 and 4 increased significantly [p<0.05]. Propanal, as the major volatile product formed during liposomal preparations, amounts from 81.2 +/- 4.1 to 118.8 +/- 2.3 microg/Kg. The differential scanning calorimetry [DSC] study showed that DHA and EPA influence the phase transition temperature of small unilamellar vesicles [SUVs] of dipalmitoyl phosphatidyl choline [DPPC]. Transmission electron microscopy [TEM] images of liposomes stained with uranyl acetate showed that the liposomes were spherical in shape and maintain high structural integrity. In conclusion, probe ultrasound of pre-formed liposomes facilitates significant loading of DHA and EPA into the nanoliposomal membrane