Asymmetric flow field‐flow fractionation of liposomes: optimization of fractionation variables

Abstract

Original Paper Asymmetric flow field-flow fractionation of liposomes: optimization of fractionation variablesThe purpose of this study was to investigate the influence of ionic strength of the carrier liquid, cross flow rate, focus flow rate, and sample load on the retention behavior of liposomes in asymmetric flow field-flow fractionation (AF4). Two differently prepared samples of large unilamellar vesicles (LUV) were used. Experiments were performed varying the factors systematically and evaluating their effect on both retention behavior of the liposomes and on particle size as obtained from online coupled multi-angle light scattering (MALS) analysis. The results showed that the focus flow rate had the least influence on the elution of liposomes. Elution of LUV is mainly governed by the chosen cross flow condition and ionic strength of the carrier liquid as well as its sample load. Optimal fractionation and size analysis were achieved using a sample load of about 10 lg, a cross flow gradient from 1.0 to 0.1 mL/min over 35 min and a carrier solution of NaNO 3 with a concentration of 10 mM. IntroductionLiposomes are vesicles consisting of an aqueous core surrounded by a phospholipid bilayer [1]. They have the ability to incorporate both amphiphilic and lipophilic compounds in their membrane-like bilayer as well as to encapsulate hydrophilic compounds [2]. Due to this ability liposomes have contributed to improving drug delivery of e.g., anthracyclines and play an important role in drug delivery research. It is widely accepted that liposomes as drug carriers may improve drug delivery in terms of therapeutic activity and safety [3 -5]. The size distribution of liposomes is important in terms of both the systemic circulation time upon injection into the blood stream and targeting abilities [3,6,7]. A study of the biodistribution of liposomes showed that the average size of the samples does not supply enough information and rather the size distribution should be used, especially for less homogeneous samples [7]. Asymmetric flow FFF (AF4) in combination with multi-angle light scattering (MALS) is the mostly applied sub-class of the FFF family and has gained increasing importance in the field of protein and peptide therapeutics as well as natural and synthetic polymers in recent years [8,9]. However, the number of publications about FFF of liposomes is rather limited compared to other applications and the technique has still not found widespread use despite it has various promising advantages over current particle size analysis methods such as photon correlation spectroscopy (PCS), and size exclusion chromatography [10,11]. For liposomal drug carriers a reliable size determination over the whole particle size spectrum from a few nanometers up to a micrometer could not be achieved with a single method so far. Such information on the other hand is essential for stringent control of the particle size which is governing the biodistribution and tumor targeting [7]. Despite a handful of successful attempts to characterize ...

Asymmetric flow field‐flow fractionation of liposomes: 2. Concentration detection and adsorptive loss phenomena

“…In an earlier study salt concentration of the carrier, sample load, crossflow rate and focusing conditions were found being critical factors for separation of liposomes and are thus chosen to assure optimum fractionation and detection of the liposome samples as performed here [28].…”

“…Focusing and fractionation conditions were as recently established [28]. Focusing was done for 7 min at a focus flow rate of 1.0 mL/min.…”

“…In a previous study the influence of crossflow rate, focusing conditions, sample load and salt concentration of the carrier onto the fractionation of liposomes in AF4 were shown [28].…”

Synaptic vesicles studied by dynamic light scattering

“…AFFF has been used for characterization and size fractionation of vesicles [22][23][24][25][26], including applications to biological vesicles [27,28]. The dispersed particles travel along a channel with a parabolic velocity profile in the principal flow direction.…”

Advanced Analytical Techniques for the Measurement of Nanomaterials in Food and Agricultural Samples: A Review

“…This combination can be ideal for measuring NMs in agricultural soil, aquatic samples, and/or humic substances containing Fe 3 O 4 /hydroxide. AF4, in combination with MALS, has gained importance in the field of food science to detect submicron size particles (Hupfeld et al, 2009).…”