Particle size determination of a submicron-sized emulsion

Abstract

The particle size distribution of a submicron-sized soybean oil-in-water emulsion stabilized by fractionated egg phospholipids was studied. Special attention was focused on the lower end of the size distribution range since a structural analysis pointed to the existence of small unilamellar vesicles (SUVs) representing an excess of emulsifier. Moreover, 31P NMR data indicated the existence of real emulsion droplets with diameters below about 100 nm. The analysis of the unfractionated emulsion by different techniques yielded the following data. The mean particle size by number was determined by photon correlation spectroscopy (PCS) under a fixed angle of 90° to be 395 nm without the Mie correction or 371 nm applying the Mie correction. Multiangle analysis resulted in a mean size of about 277–320 nm and a bimodal size distribution. The PCS measurements did not show any hint of the existence of small emulsion droplets or SUVs with diameters below 140 nm. However, more than 67% of all measured fractures on transmission electron microscopy (TEM) micrographs had diameters below 140 nm and a bimodal fracture size distribution by number was obtained. The mean fracture diameter by number was found to be 117 nm. A three-dimensional analysis of the fracture data resulted in a bimodal particle size distribution by number and a mean particle diameter of approximately 70–80 nm was calculated by two independent methods. According to the TEM results, the majority of all particles by number (SUVs and real emulsion droplets) were overlooked in the PCS experiments and only larger real emulsion droplets were included in the PCS analysis. Even though quantitative 31P NMR spectroscopy is not a conventional sizing technique, the derived cumulative frequency value of phospholipids can be related to an estimate of particle numbers. Quantitative 13P NMR data also indicated that the majority of particles by number (much more than 50%) had diameters below about 100 nm and that the system contained large numbers of emulsion droplets with diameters below about 100 nm beside SUVs. The results from TEM investigations and NMR spectroscopy did not correspond to the PCS data obtained for the native system. However, PCS measurements on ultracentrifugation fractions confirmed the existence of particles with diameters far below about 100 nm. Even though PCS has often been favoured for the determination of the size distributions of emulsions for parenteral administration (i.v. emulsions) and has been recommended for fast and reliable measurements on i.v. emulsions, its value for the determination of the complete particle size distribution of complex systems with extremely broad size distributions, such as i.v. emulsions, seems rather limited. SUVs and real emulsion droplets with diameters below about 140 nm, representing considerably more than 67% of all particles by number, were overlooked in the PCS experiments.