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PubMed1998

Vesicle size distributions measured by flow field-flow fractionation coupled with multiangle light scattering.

The separation method, flow field-flow fractionation (flow FFF), is coupled on-line with multiangle laser light scattering (MALLS) for simultaneous measurement of the size and concentration of vesicles eluting continuously from the fractionator. These size and concentration data, gathered as a function of elution time, may be used to construct both number- and mass-weighted vesicle size distributions. Unlike most competing, noninvasive methods, this flow FFF/MALLS technique enables measurement of vesicle size distributions without a separate refractive index detector, calibration using particle size standards, or prior assumptions about the shape of the size distribution. Experimentally measured size distributions of vesicles formed by extrusion and detergent removal are non-Gaussian and are fit well by the Weibull distribution. Flow FFF/MALLS reveals that both the extrusion and detergent dialysis vesicle formation methods can yield nearly size monodisperse populations with standard deviations of approximately 8% about the mean diameter. In contrast to the rather low resolution of dynamic light scattering in analyzing bimodal systems, flow FFF/MALLS is shown to resolve vesicle subpopulations that differ by much less than a factor of two in mean size.

Chemistry, Physical↗
PubMed1991

Hydrodynamic relaxation in flow field-flow fractionation using both split and frit inlets.

Two means are described for achieving hydrodynamic relaxation and thus avoiding the stopflow injection procedure in field-flow fractionation (FFF): split flow injection and frit inlet injection. The advantages, disadvantages, and the theoretical basis of these procedures are discussed. Incremental band broadening due to the final relaxation step is examined theoretically and shown to be negligible when the flow rate of the sample inlet substream is small compared to the total channel flow rate. The optimization of the sample inlet flow rate is discussed. Experimental results for both injection procedures are reported for flow/steric (or hyperlayer) FFF applied to latex standards, confirming the expected trends. However, closer examination shows that the observed incremental band broadening associated with hydrodynamic relaxation is somewhat larger than the value predicted.

Chemical Phenomena↗
PubMed2002

Size characterization of incinerator fly ash using sedimentation/steric field-flow fractionation.

Fly ash particles emitted from municipal solid waste-incinerators are of environmental concern. This study aims to investigate the applicability of sedimentation/steric field-flow fractionation (Sd/StFFF) and to develop a Sd/StFFF method for the separation and size characterization of incinerator fly ash. This study focuses on the fly ash particles larger than approxiamtely 1 microm, which comprise more than 90% (w/w) of the fly ash. Fly ash is a complex mixture of particles having various chemical compositions, sizes, shapes, and densities. Prior to Sd/StFFF analysis, fly ash particles are prefractionated into six density classes using a modified centrifugal procedure. It was found that fly ash particles are most abundant in the density range between 2.4 and 2.8 g/cm3. Different density fractions seem to contain particles of different chemical compositions. The Sd/StFFF conditions for the size-characterization of fly ash are sample concentration, approximately 0.3% (w/v); dispersing medium, 50% ethanol in water; and carrier liquid, water with 1.0% FL-70 (ionic strength approximately 0.012 M). Sd/StFFF data show no significant differences in size distribution among different density fractions. Generally, the sizes obtained from Sd/StFFF are larger than those obtained from a Coulter Multisizer and microscopy, probably because of the irregular shapes of the fly ash particles.

Air Pollutants, Occupational↗
PubMed2005

Repeatability and reproducibility of thermal field-flow fractionation in molecular weight determination of processed natural rubber.

The purpose of this study is (1) to determine the repeatability and reproducibility of thermal field-flow fractionation (ThFFF) in measuring the molecular weight of compounded natural rubber, and (2) to examine the correlation between the molecular weights obtained from ThFFF and the rheological data. 8 batches of compounded natural rubber were obtained from a thermo-mechanical mastication process, and were analyzed by ThFFF in a designed testing sequence. ThFFF analysis showed the compounded natural rubbers range in weight-average molecular weight (M(w)) from 143,000 to 360,000. By taking into account both the short term variability (repeatability) as well as the long term variability (reproducibility) of the instrument, ThFFF was shown to be able to distinguish between samples differing by as little as 21,000 in M(w) and 15,600 in number-average molecular weight, M(n) (based on cis-polyisoprene calibration); and thus is a useful tool for the molecular weight analysis of natural rubber-related materials. It was also found that the rheological data (G' and tan delta) measured on both the virgin and the compounded natural rubber correlated well with the molecular weights obtained from ThFFF when normalized.

Chemical Fractionation↗
PubMed2001

Retention behaviour of amylopectins in asymmetrical flow field-flow fractionation studied by multi-angle light scattering detection.

Asymmetrical flow field-flow fractionation (FFF) with multi-angle light scattering (MALS) detection was applied for the fractionation of amylopectins from four different sources. Samples originated from genetically modified potatoes and waxy maize. Amylopectins were dissolved in a 1 mol l(-1) sodium hydroxide solution or water. With an injected mass of 0.2 microg, well below overloading conditions, a decrease of the apparent hydrodynamic radius with increasing inlet flow-rate was observed. Moreover, a decrease of the radius of gyration with increasing elution volume was recorded by the MALS detector. Steric/hyperlayer effects are a feasible explanation for this behaviour. The observed radius of gyration at the steric inversion point was in the order of 0.3 microm, which is smaller than the theoretically calculated inversion point. Apparently, the amylopectin behave as macromolecules with a larger hydrodynamic radius than expected on basis of their radius of gyration and are subjected to significant lift forces. The results were confirmed by four fractionations with varying flow-rates but constant ratio of cross to outlet-flow. In contrast to the normal mode operation, the retention of the amylopectins depended strongly on the applied flow-rates and was close to that of a much smaller 10 kDa dextran. Apparent molar masses in the order of between 10(7) and 10(9) g mol(-1) were obtained. The results are contrasted with enzymatically degraded and oxidised starch samples that were fractionated in the normal mode.

Amylopectin↗
PubMed1993

Separation and size distribution of red blood cells of diverse size, shape, and origin by flow/hyperlayer field-flow fractionation.

Red blood cells (RBCs) from human, equine, canine, feline, and bovine whole blood samples have been separated and characterized by high-speed flow/hyperlayer field-flow fractionation (Fl/HyFFF). The elution-based separation of RBCs by this method is based mainly on the size and shape of the cell particles. The typical separation time for RBCs is less than 3 min. Size distributions can be derived from the fractograms of cell samples using a calibration plot based on retention data for uniform polystyrene beads. The method is shown to be effective both to separate and to characterize cell populations, including those with cells of abnormal shape and size. In order to investigate differences in cell morphology, shape and size changes induced by 500,000 Da Dextran were monitored. The changes in the Fl/HyFFF elution profiles indicate that the RBCs decrease in size but become partially aggregated in the presence of the dextran. These changes were found to depend on polymer concentration and specific blood samples. Some of the results from Fl/HyFFF were compared with those from the Coulter counter and from microscopy.

Animals↗
PubMed1995

Simulation of fractograms of fat emulsions in power-programmed sedimentation field-flow fractionation (SdFFF).

A quasi-empirical approach to the simulation of fractograms was examined to verify that the elution behavior of emulsions in power-based field programmed sedimentation field-flow fractionation (SdFFF) is consistent and predictable. The approach was applied to Intralipid, a commercial soybean emulsion and to an investigational medium chain triglyceride emulsion. The simulations predicted the fractograms that were obtained under various conditions of field strength, field decay and velocity of the suspending fluid, using distribution parameters obtained from one preliminary measurement of size distribution profile. Predicted fractograms were compared to experimental ones, under various fractionating powers. Good agreement was observed in most cases, in which interference of the secondary relaxation effects was not effective. The agreement confirmed the applicability of the approach to emulsions and that the simulations can be used instead of actual experiments for the optimization of their characterization by power-programmed SdFFF.

Chemical Fractionation↗
PubMed2002

Size separation of supermicrometer particles in asymmetrical flow field-flow fractionation. Flow conditions for rapid elution.

The performance of lift-hyperlayer asymmetrical flow field-flow fractionation using rapid elution conditions was tested through the separation of standard polystyrene latex particles of diameters from 2 to 20 microm. Optimization of flowrates was studied not only in order to obtain efficient and rapid separation, but also to work under conditions of various shape and steepness of the axial flow velocity gradient. Using extreme flow conditions, the five widely spaced particle sizes, 20.5-, 15.0-, 9.7-, 5.0-, and 2.0-microm diameter, could be resolved in 6 min, whereas for the narrower size range of 20.5-5.0 microm, 1 min was enough. The size selectivity in the size range 9.7-2.0 microm was studied as a function of flowrates and particle size and was found to be constant. A particle trapping device made it possible to separate particles of sizes > 10 microm, which has previously proven to be difficult in asymmetrical channels.

Chemical Fractionation↗
PubMed2003

Retention behavior of metal particle dispersions in aqueous and nonaqueous carriers in thermal field-flow fractionation.

Until quite recently, theories on thermophoresis of particles predicted very low thermophoretic velocities of metal particles in liquids. This prediction was based on the understanding that the very high thermal conductivities of metals relative to most liquid media resulted in quite low temperature gradients across the metal particle thereby leading to low net force on the particle. In this paper, we report the retention behavior of submicrometer size metal particles of silver (Ag), gold (Au), palladium (Pd) and platinum (Pt) suspended in both aqueous and organic (specifically, acetonitrile and tetrahydrofuran) carrier liquids in thermal field-flow fractionation (ThFFF). The dependence of the metal particle retention on various factors such as particle composition, amount of added electrolyte, carrier liquid composition, field strength, channel thickness, and carrier flow-rate is evaluated and discussed. A comparison in particle retention behavior among equal-sized metal, latex and silica particles is also provided.

Fractionation, Field Flow↗
PubMed1996

High-speed particle separation and steric inversion in thin flow field-flow fractionation channels.

The performance of thin (down to 71 microns thick) flow field-flow fractionation (flow FFF) channels was examined at both high channel flow-rates (up to approximately 47 ml/min) and cross flow-rates (up to approximately 11 ml/min). High levels of retention were observed, suggestive of good performance characteristics. Supporting this expectation, four sizes (0.04-0.30 microns) of polystyrene latex microbeads were baseline separated in under 3 min. Nonetheless, a plate height analysis showed that performance was still less than theoretically expected. Fast steric-hyperlayer separations of larger latex beads were observed in the same systems. Furthermore, it was shown that the steric inversion diameter was shifted down to approximately 0.23 micron thus expanding the size range to which this FFF mode is applicable. The steric inversion phenomenon observed using narrow latex standards was shown to be consistent with that found for a polydisperse polyvinylchloride latex.

Chemical Fractionation↗
PubMed2002

Characterisation of core-shell latexes by flow field-flow fractionation with multi-angle light scattering detection.

Flat channel asymmetrical flow field-flow fractionation with multi-angle light scattering (MALS) detection was used to study the swelling behaviour of core-shell particles with either carboxylated or hydroxylated shells as a function of pH and ionic strength. The equilibration time of the most heavily carboxylated core-shells appeared to be of the order of several hours. At low ionic strength (5 mM), the carboxylated core-shells showed a definite swelling response to a change in pH in the range from 5 to 10, ranging from a hydrodynamic radius increase of 24 to 118%, depending on the degree of carboxylation. A much milder response was found for the change of the root mean square (r.m.s.) radius as measured with MALS, indicating that the scattering plane is moving inwards during the swelling process due to a decreasing density of the shell. The hydroxylated core-shells appeared to be inert to a change in pH. Also the response of two expanded (pH 10) carboxylated core-shells on increasing ionic strength was studied. Comparison of the results of these ionic strength experiments with theoretical predictions based on Donnan equilibrium led us to the conclusion that a significant amount of counter-ion condensation may take place in the shells.

Chemical Fractionation↗
PubMed1999

Automated frit inlet/frit outlet flow field-flow fractionation for protein characterization with emphasis on polymeric wheat proteins.

A flow field-flow fractionation (FFF) unit fitted with a 254-nm spacer, frit inlet (FI), and frit outlet (FO) was automated for protein analysis by addition of a system controller, autosampler, and computer software to control pumps, sample loading, and data capture. Standard molecular size marker proteins and polymeric wheat storage protein extracts were used to assess the performance of the automated unit. Optimum resolution for these proteins was obtained with a sample inlet flow of 0.2 mL/min, a frit inlet flow (recirculating) of 1.4 mL/min, and a cross-flow (recirculating) of 5 mL/min using 0.05 M acetic acid containing 0.002% FL-70 as a carrier. Use of the FIFO FFF eliminates the requirement for stop-flow relaxation and pressure balancing, results in better reproducibility, and generates a 7-10-fold increase in sensitivity at the detector by concentrating fractions eluting from the channel. These improvements resulted in superior resolution of polymeric wheat protein fractions compared to those obtained previously using a standard channel with manual load and stop-flow relaxation, allowing accurate integration of peak or size range areas. Automation of this system allows unattended sample fractionation and hence markedly increases potential for sample throughput.

Chemical Fractionation↗
PubMed2001

Analysis of self-assembled cationic lipid-DNA gene carrier complexes using flow field-flow fractionation and light scattering.

Self-assembled cationic lipid-DNA complexes have shown an ability to facilitate the delivery of heterologous DNA across outer cell membranes and nuclear membranes (transfection) for gene therapy applications. While the size of the complex and the surface charge (which is a function of the lipid-to-DNA mass ratio) are important factors that determine transfection efficiency, lipid-DNA complex preparations are heterogeneous with respect to particle size and net charge. This heterogeneity contributes to the low transfection efficiency and instability of cationic lipid-DNA vectors. Efforts to define structure-activity relations and stable vector populations have been hampered by the lack of analytical techniques that can separate this type of particle and analyze both the physical characteristics and biological activity of the resulting fractions. In this study, we investigated the feasibility of flow field-flow fractionation (flow FFF) to separate cationic lipid-DNA complexes prepared at various lipid-DNA ratios. The compatibility of the lipid-DNA particles with several combinations of FFF carrier liquids and channel membranes was assessed. In addition, changes in elution profiles (or size distributions) were monitored as a function of time using on-line ultraviolet, multiangle light scattering, and refractive index detectors. Multiangle light scattering detected the formation of particle aggregates during storage, which were not observed with the other detectors. In comparison to population-averaged techniques, such as photon correlation spectroscopy, flow FFF allows a detailed examination of subtle changes in the physical properties of nonviral vectors and provides a basis for the definition of structure-activity relations for this novel class of pharmaceutical agents.

DNA↗
PubMed1997

Entropic Contribution to the Retention of Nonspherical Particles in Field-Flow Fractionation

In this paper the effect of particle shape on the entropy of nonspherical particles adjacent to a plane surface is considered. The subsequent influence on particle retention in field-flow fractionation (FFF) has been estimated. New retention equations for thin rod and disc shaped particles have been derived to cover this steric-entropic region of FFF. As particle size increases relative to the mean cloud thickness, the retention ratio for nonspherical particles is predicted to increase compared to small spherical particles of the same mass. This could result in a significant underestimation of the calculated equivalent spherical diameter (d) by FFF methods. The steric-entropic FFF equations could be used to calculate accurate d values if the large particle dimension is estimated independently (e.g., by microscopy). Alternatively, run conditions could be designed to minimize steric-entropic perturbations to the ideal retention equation.

Journal Article↗
PubMed2002

Separation and characterization of single-walled and multiwalled carbon nanotubes by using flow field-flow fractionation.

Oxidatively shortened single-walled and multiwalled carbon nanotubes were characterized by using flow field-flow fractionation (FIFFF) under normal and steric modes, respectively. Narrow size fractions were collected from FlFFF separations. The carbon nanotubes in each fraction were further characterized by using scanning and transmission electron microscopy. FlFFF separates carbon nanotubes principally on the basis of length, leading to fractions with relatively uniform lengths. The relationship between nanotube length and FlFFF elution volume is discussed.

Journal Article↗
PubMed2005

Studying protein aggregation by programmed flow field-flow fractionation using ceramic hollow fibers.

Ceramic hollow fibers have been used as separation channels for flow field-flow fractionation. The fibers were made of alpha-alumina, with a gamma-alumina layer on the inside wall acting as a semipermeable (ultrafiltration) membrane. The fibers and the separation system were tested by determining the diffusion coefficients of a series of standard proteins under various experimental conditions. Even for the smallest protein studied, a complete recovery from the fiber was obtained. A single fiber could be used for several months without problems such as leakage or fouling. The precision of the diffusion coefficient measurements was in the order of 5-10%. A good agreement with literature data was found. Programming of the cross-flow, with a time-delayed exponential decay program, was applied to extend the accessible size range for the sample components. With flow programming, the observed retention times increased linearly with the logarithm of the molar mass of proteins and aggregates, as predicted by theory. Heat-induced aggregation of beta-lactoglobulin (beta-LG) in aqueous solution was studied with the system. Upon heating, not only the extent of aggregation but also the size of the beta-LG aggregates was found to increase with the original concentration of beta-LG in solution and with the heating time. After heating in the presence of salt, very large aggregates were formed, with molar masses over 100 million. A multiangle light scattering detector was used to estimate molar masses and sizes of the protein aggregates. From the relation between the apparent diffusion coefficients and the molar masses of the aggregates, as well as from the ratio of the rms (scattering) and the hydrodyamic radii, it was concluded that the larger beta-LG aggregates behave as flexible chains in solution.

Ceramics↗
PubMed2000

Detection of nanocolloids with flow-field flow fractionation and laser-induced breakdown detection.

A new nanosize colloid detection method comprised of flow-field flow fractionation (FFFF) with laser-induced breakdown detection (LIBD) is presented, which is capable of characterizing the colloid size distribution as well as determining the number density of each size fraction in very low concentrations. The method facilitates the detection of aquatic colloids particularly in the lower range of nanometer size (< 50 nm) with the sensitivity much higher than a laser light-scattering method (LLS), i.e., the lower ppb range. The method is tested with a mixture of polystyrene colloids in three different nominal sizes, 19, 50, and 102 nm, and the results are compared with those of the LLS method. For colloids of 19-nm diameter, the present method demonstrates the detection sensitivity over 3 orders of magnitude better than that of the LLS method. The limitation of the detection sensitivity arises from the surface bleeding of a ceramic frit overlying the separation channel of the used FFFF instrument.

Chemical Fractionation↗