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The unique properties of resistant starch (RS) have made it applicable in the formulation of a broad range of functional foods. The physicochemical properties of RS play a crucial role in its applications. Recently, flow field-flow fractionation (FlFFF) has attracted increasing interest in the separation and characterization of different categories of RS. In this review, an overview of the theory behind FlFFF is introduced, and the controllable factors, including FlFFF channel design, sample separation conditions, and the choice of detector, are discussed in detail. Furthermore, the applications of FlFFF for the separation and characterization of RS at both the granule and molecule levels are critically reviewed. The aim of this review is to equip readers with a fundamental understanding of the theoretical principle of FlFFF and to highlight the potential for expanding the application of RS through the valuable insights gained from FlFFF coupled with multidetector analysis.

As located in one of the oldest and driest deserts on Earth, soils in the Atacama Desert are greatly affected by atmospheric dust deposited on soil surface and the related fate of water-dispersible colloids (WDCs,

Humic acid (HA) and reference natural organic matter (NOM) have been widely used in environmental assessment, biogeochemistry, and ecotoxicity studies. Nevertheless, similarities and differences among the commonly used model/reference NOMs and bulk dissolved organic matter (DOM) have rarely been systematically evaluated. In this study, HA, SNOM (Suwannee River NOM) and MNOM (Mississippi River NOM), both from International Humic Substances Society, and freshly collected unfractionated NOM (FNOM) were concurrently characterized to evaluate their heterogeneous nature and size-dependent chemical properties. We found that molecular weight distributions, PARAFAC-derived fluorescent components, and size-dependent optical properties are NOM-specific and highly variable with pH. The < 1 kDa DOM abundance followed the order of HA < SNOM < MNOM < FNOM. In addition, FNOM was more hydrophilic and contained more protein-like and autochthonous components with a higher UV-absorbance ratio index (URI) and biological fluorescence index, whereas HA and SNOM contained more allochthonous, humic-like components with a higher aromaticity and lower URI. Significant differences in molecular composition and size spectra between FNOM and model/reference NOMs suggest that environmental role of NOMs should be evaluated at the levels of molecular weight and functionalities under the same experimental conditions and that HA and SNOM may not represent bulk NOM in the environment. This study provides new information about similarities and differences in DOM size-spectra and chemical properties between reference NOMs and in-situ NOM and highlights the need to better understand the heterogenous roles of NOMs in regulating the toxicity/bioavailability and environmental fate of pollutants in aquatic environments. [ABSTRACT FROM AUTHOR]

Due to the increased applications of engineered elemental metal nanoparticles (EMNPs) in recent years, increased attention has been devoted to their release into the environment. EMNPs pose potential risks to living organisms, including human beings. Hence, the characterization of EMNPs in the environment has gained significant importance. Among the various techniques reported for the characterization of EMNPs, on-line coupling of flow field-flow fractionation with inductively coupled plasma mass spectrometry (F4-ICPMS) has been well established for the simultaneous separation, identification, and quantification of EMNPs, especially in the complex matrices of environmental samples. Thus, this review focuses on the specific advantages of the F4 method, especially the asymmetrical F4 (AF4) and hollow fiber F4 (HF5) methods, in the separation of EMNPs, the general development of AF4-ICPMS and HF5-ICPMS techniques, and recent advances in the application of these hyphenated techniques in examining the occurrence and transformation of EMNPs in the environment. Finally, several perspectives on these techniques have been put forward. [ABSTRACT FROM AUTHOR]

• Field programming in flow FFF and a thickness-tapered channel were compared. • Evaluation was made with the separation of submicron PS particles. • Field programming assures a higher resolution but tapered channel offers a faster separation. • Tapered channel resolves larger particles better than a conventional channel. The thickness-tapered channel structure in flow field-flow fractionation (FlFFF), recently introduced by constructing a channel with a linear decrease in thickness along its length, demonstrated effectiveness in steric/hyperlayer separation of supramicron particles with improvements in separation speed, elution recovery, and an expanded dynamic size range of separation. In this study, we conducted a comparative analysis of the performance between the impact of field (or crossflow rate) programming or outflow rate programming for the separation of polystyrene latex standards (50 ∼ 800 nm) with a conventional channel having uniform thickness and a thickness-tapered channel without programming. Outlet flow rate and crossflow rate conditions were also varied. Although the particle size resolution of the tapered channel does not surpass that of field programming in uniform thickness channel, it achieves higher-speed separation without a significant loss of resolution and without the need for a complex flow controller system even at a low outflow rate condition. Furthermore, it yielded an improved resolution for particles close to the steric transition regime (400 ∼ 600 nm) in the normal mode of separation. Due to the continuous increase in mean flow velocity down the channel, the tapered channel exhibits flexibility in separating submicron-sized particles at high crossflow rate conditions or low outflow rate conditions, of which the latter can be advantageous when coupled with mass spectrometry in a miniaturized setup. [ABSTRACT FROM AUTHOR]

Abstract Flow field-flow fractionation (Fl-FFF) for silica nanoparticles with offline inductively coupled plasma mass spectrometry (ICP-MS) was applied to investigate the adsorption behavior of tin onto silica nanoparticles. Effect of carrier solutions and membranes was studied to achieve better separation for silica nanoparticles prior to tin detection using ICP-MS. Investigation was carried out by using 0.25 mM ammonium carbonate and 0.02% FL-70 with 0.02% NaN3 as carrier solutions with 1 kDa regenerated cellulose (RC), 10 kDa regenerated cellulose (RC), and 10 kDa polyethersulfone (PES) membranes. Ammonium carbonate carrier solution with suitable ionic strength provided good separation with minimization of particle-membrane interaction. Better retention was shown by employing 10 kDa RC membrane. Furthermore, Fl-FFF was employed for the separation of silica nanoparticles incubated with tin. Fractions eluted from Fl-FFF were collected and then introduced into ICP-MS. Tin was adsorbed onto silica nanoparticles with different adsorption capabilities depending on particle size. Adsorption of tin was greater on the smaller size of silica nanoparticles compared to the bigger size with the adsorption percentage of 98.5, 44.9, and 6.5 for 60 nm, 100 nm, and 200 nm, respectively. Size-dependent adsorption of tin was in good agreement with surface area per volume of silica nanoparticles.

Water-dispersible colloids (WDCs) are vital for trace element migration, but there is limited information about the abundance, size distribution and elemental composition of WDC-bound thallium (Tl) and arsenic (As) in mining-contaminated soils and sediments solutions. Here, we investigated the potential mobilization of WDC-bound Tl and As in soils and sediments in a typical Tl/As-contaminated area. Ultrafiltration results revealed on average > 60% of Tl and As in soil solution (< 220 nm) coexisted in colloidal form whereas Tl and As in sediment solution primarily existed in the truly dissolved state (< 10 kDa) due to increased acidity. Using AF4-UV-ICP-MS and STEM-EDS, we identified Fe-bearing WDCs in association with aluminosilicate minerals and organic matter were main carriers of Tl and As. SAED further verified jarosite nanoparticles were important components of soil WDC, directly participating in the migration of Tl and As. Notably, high pollution levels and solution pH promoted the release of Tl/As-containing WDCs. This study provides quantitative and visual insights into the distribution of Tl and As in WDC, highlighting the important roles of Fe-bearing WDC, soil solution pH and pollution level in the potential mobilization of Tl and As in contaminated soils and sediments. [Display omitted] • > 60% of Tl and As in soil solution occurred in water-dispersible colloids (WDCs). • Tl/As-bearing WDCs comprised Fe oxides, aluminosilicates and OM. • Jarosite was an important component of WDCs involved in Tl/As migration. • High pollution levels and solution pH promoted the release of Tl/As-bearing WDCs. [ABSTRACT FROM AUTHOR]

Increasing amounts of engineered nanomaterials such as TiO2 and CeO2 are released into air, waters, soils, and sediments. However, assessing the human-made origin of those nanomaterials is rather difficult because Ti- and Ce-rich particles are naturally present in soils and sediments at concentrations typically much higher than estimated concentrations of engineered nanomaterials. In addition, analysis is complicated by the interactions and aggregation of nanoparticles with environmental particles. Therefore, more knowledge on the properties of natural nanomaterials is needed to distinguish engineered nanomaterials in natural systems. Here, we extracted soil nanomaterials with six extractants and compared recovery and disaggregation to primary particles. Nanomaterials were characterized for hydrodynamic diameter and zeta potential by dynamic light scattering, size-based elemental distribution by field-flow fractionation coupled with inductively coupled plasma-mass spectroscopy, and morphology by transmission electron microscopy. Results show that nanomaterial concentrations increased from CH3COOH–NaCl–water (lowest), to water or NaCl–water, Na2CO3, Na4P2O7, and NaCl–Na4P2O7 (highest). Na4P2O7 was the most efficient extractant that induced the release of primary nanomaterials from microaggregates. Although sodium carbonate extracted relatively high concentrations of nanomaterials, the extracted nanomaterials occurred mainly as aggregates of primary nanomaterials. Ultrapure water, sodium chloride and acetic acid resulted in poor nanomaterial extraction and broad size distributions. Elemental ratios illustrate that Ti is associated with Nb, Ta, and V, and that Ce is associated with rare earth elements such as La, Eu, Y, Ho, Er, Tm, and Yb. Our findings indicate that size, size distribution, and elemental ratios can be used as fingerprints to differentiate engineered nanomaterials such as TiO2 and CeO2 from natural nanomaterials in complex media. [ABSTRACT FROM AUTHOR]

We investigated the effect of oxidative stress (OS) on lipidomic perturbations in the subcellular fractions and exosomes of human embryonic kidney (HEK) 293 cells using asymmetrical flow field–flow fractionation (AF4) and nanoflow ultrahigh performance liquid chromatography–electrospray ionization–tandem mass spectrometry (nUHPLC-ESI-MS/MS). We treated HEK 293 cells with hydrogen peroxide (H 2 O 2) and fractionated the cell lysates using AF4 to determine the change in size and population of the subcellular fractions and exosomes, and to obtain narrow size fractions for lipid analysis. A total of 438 lipids from 642 identified species—including oxidized lipids—were quantified. The relative amount of secreted exosomes increased by 28% during OS, whereas the amount of subcellular species decreased by 35%. There was a significant increase in the level of oxidized phospholipids in the mitochondrion-enriched subcellular fractions, but not in the exosomes. Most high-abundance triacylglycerol (TG) species increased in the stressed cells, whereas they decreased in the exosomes. During OS, ceramides involved in the apoptotic mitochondrial pathway were accumulated in the subcellular fractions, whereas their levels were unaffected in the exosomes. The present study demonstrated that AF4 and nUHPLC-ESI-MS/MS can be used to investigate lipid alterations in subcellular and extracellular species during OS, and the pathological relationships in diseases caused by reactive oxygen species. Image 1 • Oxidative stress effect on lipids of subcellular fractions and exosome is investigated. • Size and population changes in subcellular fractions and exosome are examined by flow FFF. • Oxidized phospholipids, TG, and Cer in subcellular fractions and exosome showed different patterns of alteration. • Flow FFF and nUHPLC-ESI-MS/MS can be a powerful method to analyze lipidomic perturbation in cellular components. [ABSTRACT FROM AUTHOR]

Two-line ferrihydrite (2LFh) was aged for 12 years under ambient conditions and sheltered from light in the presence of Lu(III) used as surrogate for trivalent actinides. 2LFh aging produced hematite rhombohedra with overgrown acicular goethite particles. Analysis of the homogeneous suspension by asymmetrical flow field-flow fractionation (AsFlFFF) coupled to ICP-MS indicated that particles have a mean hydrodynamic diameter of about 140 nm and the strong correlation of the Fe and Lu fractograms hinted at a structural association of the lanthanide with the solid phase(s). Unfortunately, recoveries were low and thus results cannot be considered representative of the whole sample. The suspension was centrifuged and X-ray absorption spectroscopy (XAS) at the Lu L3-edge on the settled particles indicated that Lu(III) is sixfold coordinated by oxygen atoms, pointing to a retention by structural incorporation within particles. This result is consistent with AsFlFFF results on the same suspension without centrifugation. The detection of next nearest Fe and O atoms were consistent with the structure of goethite, ruling out incorporation within hematite. After centrifugation of the suspension, only nanoparticulate needle-like particles, very likely goethite, could be detected in the supernatant by ESEM. AsFlFFF data of the supernatant were comparable to that obtained for the homogeneous suspension, whereas XAS indicated that Lu(III) is predominantly present as dissolved species in the supernatant. Results from both techniques can be interpreted as a major fraction of Lu present as aqueous ions and a minor fraction as structurally incorporated. Findings from this study are corroborated by STEM-HAADF data and results from DFT calculations in a companion paper. [ABSTRACT FROM AUTHOR]

Abstract The current study documents enhanced apparent solubility of the BCS class II drug celecoxib (CXB) when formulated as solid phospholipid dispersion (SPD) with either mono- or diacyl-phospholipids by freeze drying from hydro-alcoholic solvent. The enhanced solubility upon dispersion in buffer or fasted state simulated intestinal fluid (FaSSIF) is interpreted to be due to two effects: (1) amorphization of CXB, inducing supersaturation, which is also observed when CXB is freeze dried in the absence of phospholipids and (2) association of CXB with spontaneously forming colloidal structures, such as vesicles and/or micelles, promoting solubilization. The latter effect depended on the CXB-to-phospholipid ratio, where monoacyl-phospholipid was a more efficient solubilizer than diacyl-phospholipid. In the case of diacyl-phospholipid, solubilization also depended strongly on the dispersion medium, where FaSSIF induced a more pronounced solubilization effect than buffer. In contrast, a significantly enhanced in-vitro permeability of CXB across a biomimetic barrier (Permeapad®) was found only with low lipid contents up to a CXB to phospholipid mass-ratio of 1:10 or in the absence of phospholipid; above this critical ratio, permeability was not enhanced, i.e. comparable to that observed with a suspension of non-processed (crystalline) drug. This non-linear dissolution-/permeation-behavior was observed independently of (1) the type of phospholipid (monoacyl- or diacyl-) employed and (2) the dispersion medium (buffer or FaSSIF), despite the fact that different patterns of co-existing colloidal states were observed from mono-/diacyl-phospholipid formulations in buffer/FaSSIF (small bile salt micelles, intermediate size mixed micelles and large vesicular structures), assessed by asymmetric flow field-flow fractionation/multi angle laser light scattering. A uniform mechanistic hypothesis is presented to describe the impact of phospholipids on CXB permeation behavior: Obviously, the critical drug-to-phospholipid ratio represents a compromise between optimal stabilization of the amorphous state-induced supersaturation and reduced thermodynamic activity of CXB due to association with colloidal states, where the type of colloidal state (vesicle or micelle) appears to be of minor importance. Graphical abstract Unlabelled Image [ABSTRACT FROM AUTHOR]

Abstract In the present study, we characterized the sources of dissolved organic matter (DOM) and the processes controlling the evolution of DOM in a large anthropogenically perturbed estuary in Southern China. Three-dimensional excitation emission matrices (3D EEMs) revealed that DOM in the estuary was primarily from autochthonous origin although terrestrial sources also contributed to the DOM production. Parallel factor analysis (PARAFAC) of the EEMs dataset indicated that DOM in the estuary was a legacy from the upstream wastewater discharging activities, and estuarine mixing was the primary mechanism responsible for the decreasing concentrations of DOM components. However, non-conservative behavior of DOM components was also observed along the center transect as a result of additional removal process and spatial heterogeneity in the distribution of DOM. Seasonal influence on the concentrations and compositions of colloidal DOM in the estuary was also found. Asymmetric flow field-flow fractionation (AF4) analysis showed distinct distribution patterns of the colloidal DOM during the dry and wet seasons. Behavior of colloidal DOM was size-independent and the distribution of colloidal DOM was controlled by multiple depletion and replenishment cycles in the dry season, in contrast to those observed in the wet season. By combining both EEMs-PARAFAC modeling and AF4 technique, this study provided important information on the sources and fate of bulk and colloidal DOM in a large estuary with intensive anthropogenic influences. Graphical abstract Image 1 Highlights • DOM in the estuary was primarily from autochthonous origin although terrestrial sources also contributed. • Estuarine mixing was responsible for the decrease of humic-like and protein-like DOM with salinity. • Colloidal DOM went through multiple depletion and replenishment cycles in the dry season. • Colloidal DOM was associated with both allochthonous and autochthonous origins during the wet season. [ABSTRACT FROM AUTHOR]

Abstract Knowledge of the dynamic changes in molecular size of natural colloidal organic matter (COM) along the aquatic continuum is of vital importance for a better understanding of the environmental fate and ecological role of dissolved organic matter and associated contaminants in aquatic systems. We report here the pH- and cation-dependent size variations of COMs with different sources (river and lake) quantified using flow field-flow fractionation (FIFFF), fluorescence spectroscopy and parallel factor analysis (PARAFAC), attenuated total reflectance Fourier transform infrared (ATR–FTIR) spectroscopy, and zeta potential analysis. Increasing pH caused a decline in molecular sizes and an obvious size transformation from the >10 kDa to 5–10 kDa and further to 1–5 kDa size fraction, whereas the opposite trend was observed for increasing cation (e.g., Ca2+ and Cu2+) abundance. Compared with lakewater COM, the riverwater COM exhibited a greater pH-dependent dispersion but less extent in cation-induced aggregation, demonstrating that the dispersion and aggregation dynamics were highly dependent on COM source and solution chemistry (e.g., pH and cations). Based on ATR–FTIR analysis, the extensive dissolution of C=O and C–O functional groups resulted in a greater pH-dependent dispersion for river COM. Fluorescence titration revealed that, despite their similar cation-induced aggregation behavior, the binding constants of all the PARAFAC-derived components for Cu2+ were 1–2 orders of magnitude higher than those for Ca2+ (logK M : 4.54–5.45 vs. 3.35–3.70), indicating a heterogeneous nature in cation-DOM interactions. The greater extent of decline in zeta potential for lake COM suggested a Ca-induced charge neutralization and aggregation mechanism. However, for Cu-induced aggregation, chemical complexation was the predominant pathway for the river COM, with higher binding constants, while charge neutralization and chemical complexation co-induced the aggregation of lake COM. Thus, natural COMs may have different environmental behavior along the aquatic continuum and further affect the fate and transport of contaminants in aquatic environments. Graphical abstract Image 1 Highlights • Increased pH causes COM to disperse from larger (>10 and 5–10 kDa) to smaller (1–5 kDa) sized NOM. • Increased cation enhances COM coagulation/aggregation to form larger sized COM. • River COM exhibited greater pH-dependent dispersion but less cation-induced aggregation than lake COM. • Extensive dissolution of C=O and C–O groups accounted for a greater pH-induced dispersion for river COM. • Compared to river COM, charge neutralization and chemical complexation co-induced the aggregation of lake COM. [ABSTRACT FROM AUTHOR]

Abstract A comprehensive lipid analysis was performed at the plasma lipoprotein level in patients with acute coronary syndrome (ACS) and stable coronary artery disease (CAD). Because the lipids in lipoproteins are related to the pathology of the cardiovascular system, lipoprotein-specific lipid analysis can be useful for understanding the mechanism of lipid-associated cardiovascular diseases. Lipoproteins were size-sorted into high density lipoproteins (HDL) and low density lipoproteins (LDL) using asymmetrical flow field-flow fractionation, then lipids of each lipoprotein were analysed using nanoflow ultrahigh performance liquid chromatography-electrospray ionization-tandem mass spectrometry. A total of 365 lipids were structurally identified and quantified by selected reaction monitoring method. Two high abundance lysophosphatidylcholines (16:0 and 18:0) were significantly increased only in the HDL of the ACS group (vs. the stable CAD group). Phosphatidylethanolamines (38:5 and 40:5) significantly increased in ACS by >2-fold in both lipoproteins. (18:0, 22:6)-diacylglycerol increased in ACS by 3.5-fold only in LDL; however, most high abundance triacylglycerols decreased 2-fold in both lipoproteins. The present study revealed the usefulness of lipoprotein-specific analysis of lipids in distinguishing ACS from stable CAD, and the selected lipids analysed in this study may be useful in the development of lipid markers for the early detection of ACS. [ABSTRACT FROM AUTHOR]

Flow field-flow fractionation (FlFFF) with inductively coupled plasma mass spectrometric (ICP-MS) detection was applied for estimating the hydrodynamic diameter of gold nanoparticles (AuNPs). Hydrodynamic diameters of AuNPs of the same core diameter but with different surface coatings were different because the coating agents and their properties were different. The challenge of this work is due to the fact that AuNPs with various types of surface coatings exhibited different interactions in the FlFFF channel, leading to different retention behaviors. Therefore, we are interested in finding suitable FlFFF conditions for estimating the hydrodynamic diameter of AuNPs with various types of electrostatic stabilizing agents [tannic acid (TA) and citrate (CT)] and steric stabilizing agents [polyethylene glycol (PEG), polyvinylpyrrolidone (PVP), and branched polyethylene imine (BPEI)]. Different types of carrier liquids (DI water, 0.02% FL-70, 0.05% SDS, and 30 mM Tris buffer) and membrane materials [regenerated cellulose (RC) and polyethersulfone (PES) membranes] were investigated. Generally, FlFFF was applied for size characterization of nanoparticles based on FlFFF theory but the interactions between AuNPs and membrane affected the retention and the experimentally obtained hydrodynamic diameters of AuNPs from the FlFFF system. With DI water as a carrier liquid with RC or PES membranes, the hydrodynamic diameters of negatively charged particles (TA-, CT-, PVP-, and PEG-stabilized AuNPs) from FlFFF corresponded well with the hydrodynamic diameters from dynamic light scattering (DLS). Interestingly, it was possible to estimate hydrodynamic diameters of AuNPs in the mixture by using FlFFF whereas it was not possible with the use of DLS within the size range studied. This work summarized the possible interactions between AuNPs with various coating agents and membrane materials in different carrier liquids to give guidelines on the suitable conditions of FlFFF for further applications on AuNP hydrodynamic diameter estimation. [ABSTRACT FROM AUTHOR]

Alzheimer’s disease (AD) is an irreversible neurodegenerative disorder with the clinical symptom of the progressive loss of cognitive function and mild cognitive impairment (MCI) is a translational state between cognitive changes of normal aging and AD. Lipid metabolism and pathogenesis of Alzheimer’s disease (AD) are closely linked. Despite obviously discrete lipidome constitutions across lipoproteins, lipidomic approaches of AD has been mostly conducted without considering lipoprotein-dependent alterations. This study introduces a combination of asymmetrical flow field-flow fractionation (AF4) and nanoflow ultrahigh performance liquid chromatography-tandem mass spectrometry (nUHPLC-ESI-MS/MS) for a comprehensive lipid profiling in different lipoprotein level of patients plasma with AD and amnestic MCI in comparison to age-matched healthy controls. Lipoproteins in plasma samples were size-sorted by a semi-preparative scale AF4, followed by non-targeted lipid identification and high speed targeted quantitation with nUHPLC-ESI-MS/MS. It shows 14 significantly altered high abundance lipids in AD, exhibiting >2-fold increases ( p  < 0.01) in LDL/VLDL including triacylglycerol, ceramide, phosphatidylethanolamine, and diacylglycerol. Three lipid species (triacylglycerol 50:1, diacylglycerol 18:1_18:1, and phosphatidylethanolamine 36:2) showing a strong correlation with the degree of brain atrophy were found as candidate species which can be utilized to differentiate the early stage of MCI when simple Mini-Mental State Examination results were statistically incorporated. The present study elucidated lipoprotein-dependent alterations of lipids in progression of MCI and further to AD which can be utilized for the future development of lipid biomarkers to enhance the predictability of disease progress. [ABSTRACT FROM AUTHOR]

The angular dependency of light scattering intensity from differently sized particles strongly influences the apparent particle size distribution, as determined by dynamic light scattering (DLS) methods. Manufactured nanomaterials have size distributions more or less; therefore, the effect of detecting the angular dependency of the apparent size distribution by DLS is crucial. Commercial DLS instruments typically have two different types of detector angular position. The first is a detector angled at 90°, and the other is a backscattering angle detector. We therefore investigated the coverage and angular dependency when determining the relative concentrations of nanoparticles in polystyrene latex samples with a bimodal size distribution, using DLS methods both experimentally and theoretically. We used five differently sized polystyrene latex particles (one was a 70-nm nanoparticle and the others were various submicron-sized particles) in a variety of mixtures (the ratio of the difference of particle sizes ranged from approximately 2 to 7) to investigate the coverage and angular dependency of the recognition of the relative concentration ratio. In the case of size difference of approximately a factor of 2 or 3 between the two mixed particles (one was fixed at 70 nm), for DLS measurements at light scattering detector angles ranging from 60° to 150°, the homodyne photon correlation functions were approximately straight lines for mixtures of two differently sized polystyrene latex particles. The straight homodyne photon correlation functions were caused by the relatively strong light scattering from larger submicron particles masking the weaker light scattering from the smaller nanoparticles. As a result, DLS analysis could not recognize the relative concentration of nanoparticles in the mixture. In contrast to these samples, for mixtures of two differently sized polystyrene latex particles (one was 70 nm in size) with a size difference of a factor of 5, the homodyne correlation functions displayed an obvious curve for angles larger than 120°. This curve reflected an appropriate relative concentration ratio for the two differently sized polystyrene latex particles. Furthermore, for a mixture of two differently sized particles (one was again 70 nm) with size differences of a factor of 7, the homodyne correlation functions showed a clearly curved shape for detector angles larger than 90°, and yielded appropriate relative concentration ratios for the two different sizes of polystyrene latex particles. These observations were supported by theoretical investigation using Mie theory and asymmetric flow field-flow fractionation measurements with a multi-angle light scattering detector. Our investigation is crucial for achieving some degree of concordance on the determination of the size distribution of particles using DLS methods in industrial and academic fields. [ABSTRACT FROM AUTHOR]

Asymmetric flow field-flow fractionation (AF4) coupled with UV-Vis spectroscopy, multi-angle light scattering (MALS) and refractive index (RI) detection has been applied for the characterization of MIL-100(Fe) nanoMOFs (metal-organic frameworks) loaded with nucleoside reverse transcriptase inhibitor (NRTI) drugs for the first time. Empty nanoMOFs and nanoMOFs loaded with azidothymidine derivatives with three different degrees of phosphorylation were examined: azidothymidine (AZT, native drug), azidothymidine monophosphate (AZT-MP), and azidothymidine triphosphate (AZT-TP). The particle size distribution and the stability of the nanoparticles when interacting with drugs have been determined in a time frame of 24 h. Main achievements include detection of aggregate formation in an early stage and monitoring nanoMOF morphological changes as indicators of their interaction with guest molecules. AF4-MALS proved to be a useful methodology to analyze nanoparticles engineered for drug delivery applications and gave fundamental data on their size distribution and stability.ᅟ [ABSTRACT FROM AUTHOR]

Asymmetrical flow field-flow fractionation (AF4) and hollow-fiber flow field-flow fractionation (HF5) are techniques widely used in analytical, industrial and biological analyses. The main problem in all AF4 and HF5 analyses is sample loss due to analyte-membrane interactions. In this work the impact of liquid carrier composition on latex nanoparticles (NPs) separation in water and two different concentrations of NH4NO3 was studied. In AF4, a constant trend of decreasing the size of 60 and 121.9 nm particles induced by the ionic strength of the carrier liquid has been observed. In contrast, an agglomeration effect of the biggest 356 nm particles was observed when increasing ionic strength, which induced a significant drop of recovery to 35%. H5F provides better resolution and intensified peaks of NPs, but careful optimisation of system parameters is mandatory to obtain good separation. [ABSTRACT FROM AUTHOR]

Colloidal phases (self-assemblies) in aqueous dispersions of selected binary bile salt/phospholipid blends were studied utilizing the combined analytical approach of asymmetrical flow field-flow fractionation (AF4) and multi-angle laser light scattering (MALLS) in order to resolve the co-existence of different colloidal assemblies. The binary blends were prepared by freeze-drying from tert -butanol/water co-solvent solutions. The blends contained one of two bile salts (sodium taurocholate (TC) or sodium glycodeoxycholate (GDX)) and a mono- or di-acyl phospholipid (lyso-phosphatidylcholine (L-PC) and phosphatidylcholine (PC), respectively). Bile salt and phospholipid (PL) concentrations and their respective ratios were varied systematically within the physiological range found in human intestinal fluids. Furthermore, the BCS class II drug Celecoxib was incorporated in selected blends to assess its potential impact on colloidal phases. To further investigate the smallest self-assemblies observed in AF4/MALLS analysis, dispersions of TC and GDX, respectively, were prepared and analyzed by dynamic light scattering (DLS). AF4/MALLS analysis revealed that binary bile-salt/phospholipid blends form three distinct particle fractions, when the concentration of bile-salt was sufficiently high (≥3.5 mM). Those fractions were assumed to be very small pure bile-salt dimeric/oligomeric self-assemblies (Ø ≈ 2–3 nm), mid-sized mixed micelles (Ø ≈ 10–50 nm) and large liposomes/aggregates (Ø ≈ 150–280 nm). If present, Celecoxib was found solubilized within the structures, but at the lowest TC concentration triggered the formation of an additional (vesicular) phase. [ABSTRACT FROM AUTHOR]

This study suggests a novel method for determination of the channel height in asymmetrical flow field-flow fractionation (AF4), which can be used for calibration of the channel for hydrodynamic radius determinations. The novel method uses an oil-in-water nanoemulsion together with multi angle light scattering (MALS) and elution theory to determine channel height from an AF4 experiment. The method is validated using two orthogonal methods; first, by using standard particle elution experiments and, secondly, by imaging an assembled and carrier liquid filled channel by x-ray computed tomography (XCT). It is concluded that the channel height can be determined with approximately the same accuracy as with the traditional channel height determination technique. However, the nanoemulsion method can be used under more challenging conditions than standard particles, as the nanoemulsion remains stable in a wider pH range than the previously used standard particles. Moreover, the novel method is also more cost effective. [ABSTRACT FROM AUTHOR]

Flow field-flow fractionation (FlFFF) is a gentle technique that does not require a stationary phase and allows for the non-destructive isolation/separation of biomacromolecules and bioparticulate species in biological samples. Given the large separation range (i.e., 1 nm to 100 μm), FlFFF is theoretically applicable for the full spectrum of vital biological species. We provide an overview of the basic separation mechanism and development of FlFFF, followed by discussion of the current applications of FlFFF in the characterization of representative biomacromolecules, bionanoparticles, and biomicroparticles in biological samples, particularly new advances in the last 5 years. Furthermore, the emerging applications of FlFFF in extracellular vesicle research are systematically discussed for the first time. Finally, we posit that more efforts are needed to direct the separation and characterization of biomacromolecules, bionanoparticles, and biomicroparticles using FlFFF. Overall, this review provides current status, developmental trends, and further advances of FlFFF in life science. • Full-spectrum separation of biological species by two FlFFF separation modes. • The potential of FlFFF in disease diagnosis and treatment. • Very recent use of FlFFF in the separation and characterization of EVs. • Some perspectives and future directions of FlFFF in life science. [ABSTRACT FROM AUTHOR]

Polymer self-assemblies joining oppositely charged chains, known as polyion complexes (PICs), have been formed using poly(ethyleneoxide – b – acrylic acid)/poly( l -lysine), poly(ethyleneoxide-b-acrylic acid)/dendrigraft poly( l -lysine) and poly[(3-acrylamidopropyl) trimethylammonium chloride – b – N – isopropyl acrylamide]/poly(acrylic acid). The self-assemblies have been first characterized in batch by Dynamic Light Scattering. In a second step, their analysis by Flow Field-Flow Fractionation techniques (FlFFF) was examined. They were shown to be very sensitive to shearing, especially during the focus step of the fractionation, and this led to an incompatibility with asymmetrical FlFFF. On the other hand, Frit Inlet FlFFF proved to be very efficient to observe them, either in its symmetrical (FI-FlFFF) or asymmetrical version (FI-AsFlFFF). Conditions of elution were found to optimize the sample recovery in pure water. Spherical self-assemblies were detected, with a size range between 70–400 nm depending on the polymers. Compared to batch DLS, FI-AsFlFFF clearly showed the presence of several populations in some cases. The influence of salt on poly(ethyleneoxide-b-acrylic acid) (PEO-PAA) 6000-3000/dendrigraft poly( l -lysine) (DGL 3) was also assessed in parallel in batch DLS and FI-AsFlFFF. Batch DLS revealed a first process of swelling of the self-assembly for low concentrations up to 0.8 M followed by the dissociation. FI-AsFlFFF furthermore indicated a possible ejection of DGL3 from the PIC assembly for concentrations as low as 0.2 M, which could not be observed in batch DLS. [ABSTRACT FROM AUTHOR]

Flow-Field Flow Fractionation (Flow-FFF), coupled with online detection systems is one of the most promising tools available for the separation and quantification of engineered nanoparticles (ENPs) in complex matrices. To correctly relate the retention of nanoparticles in the Flow-FFF-channel to the particle size, ideal separation conditions must be met. This requires optimization of the parameters that influence the separation behavior. The aim of this study was therefore to systematically investigate and evaluate the influence of parameters such as the carrier liquid, the cross flow, and the membrane material, on the separation behavior of two metallic ENPs. For this purpose the retention, recovery, and separation efficiency of sterically stabilized silver nanoparticles (AgNPs) and electrostatically stabilized gold nanoparticles (AuNPs), which represent two materials widely used in investigations on environmental fate and ecotoxicology, were investigated against a parameter matrix of three different cross-flow densities, four representative carrier solutions, and two membrane materials. The use of a complex mixture of buffers, ionic and non-ionic surfactants (FL-70 solution) together with a medium cross-flow density provided an acceptable compromise in peak quality and recovery for both types of ENPs. However, these separation conditions do not represent a perfect match for both particle types at the same time (maximized recovery at maximized retention). It could be shown that the behavior of particles within Flow-FFF channels cannot be predicted or explained purely in terms of electrostatic interactions. Particles were irreversibly lost under conditions where the measured zeta potentials suggested that there should have been sufficient electrostatic repulsion to ensure stabilization of the particles in the Flow-FFF channel resulting in good recoveries. The wide variations that we observed in ENP behavior under different conditions, together with the different behavior that has been reported in published literature for the same NPs under similar conditions, indicate a need for improvement in the membrane materials used for Flow-FFF analysis of NPs. This research has shown that careful adjustment of separation conditions can result in acceptable, but not ideal, separation conditions for two fundamentally different stabilized materials, and that it may not be possible to separate a set of different particles under ideal conditions for each particle type. This therefore needs to be taking into account in method development and when interpreting FFF results from complex samples. [ABSTRACT FROM AUTHOR]

Facing the lack of studies on characterization and quantification of cerium oxide nanoparticles (CeO 2 NPs), whose consumption and release is greatly increasing, this work proposes a method for their sizing and quantification by Flow Field-flow Fractionation (FFFF) coupled to Inductively Coupled Plasma-Mass Spectrometry (ICP-MS). Two modalities of FFFF (Asymmetric Flow- and Hollow Fiber-Flow Field Flow Fractionation, AF4 and HF5, respectively) are compared, and their advantages and limitations discussed. Experimental conditions (carrier composition, pH, ionic strength, crossflow and carrier flow rates) are studied in detail in terms of NP separation, recovery, and repeatability. Size characterization of CeO 2 NPs was addressed by different approaches. In the absence of feasible size standards of CeO 2 NPs, suspensions of Ag, Au, and SiO 2 NPs of known size were investigated. Ag and Au NPs failed to show a comparable behavior to that of the CeO 2 NPs, whereas the use of SiO 2 NPs provided size estimations in agreement to those predicted by the theory. The latter approach was thus used for characterizing the size of CeO 2 NPs in a commercial suspension. Results were in adequate concordance with those achieved by transmission electron microscopy, X-ray diffraction and dynamic light scattering. The quantification of CeO 2 NPs in the commercial suspension by AF4-ICP-MS required the use of a CeO 2 NPs standards, since the use of ionic cerium resulted in low recoveries (99 ± 9% vs. 73 ± 7%, respectively). A limit of detection of 0.9 μg L −1 CeO 2 corresponding to a number concentration of 1.8 × 1012 L −1 for NPs of 5 nm was achieved for an injection volume of 100 μL. [ABSTRACT FROM AUTHOR]

Filter-extrusion is a widely used technique for down-sizing of phospholipid vesicles. In order to gain a detailed insight into size and size distributions of filter-extruded vesicles composed of egg phosphatidyl-choline (with varying fractions of cholesterol) – in relation to extrusion-parameters (pore-size, number of filter passages, and flow-rate), flow field-flow fractionation in conjunction with multi-angle laser light scattering (AF4-MALLS, Wyatt Technology Corp., Santa Barbara, CA) was employed. Liposome size-distributions determined by AF4-MALLS were compared with those of dynamic light scattering and correlated with cryo-transmission electron microscopy and31P-NMR-analysis of lamellarity. Both the mean size of liposome and the width of size distribution were found to decrease with sequential extrusion through smaller pore size filters, starting at a size range of ≈70–415 nm upon repeated extrusion through 400 nm pore-filters, eventually ending with a size range from ≈30 to 85 nm upon extrusion through 30 nm pore size filters. While for small pores sizes (50 nm), increased flow rates resulted in smaller vesicles, no significant influence of flow rate on mean vesicle size was seen with larger pores. Cholesterol at increasing mol fractions up to 0.45 yielded bigger vesicles (at identical process conditions). For a cholesterol mol fraction of 0.5 in combination with small filter pore size, a bimodal size distribution was seen indicating cholesterol micro-crystallites. Finally, a protocol is suggested to prepare large (∼ 300 nm) liposomes with rather narrow size distribution, based on the filter extrusion at defined flow-rates in combination with freeze-/thaw-cycling and bench-top centrifugation. [ABSTRACT FROM PUBLISHER]

• P increased in all colloidal fractions with long-term cultivation management. • The nanoparticle fraction turned into a complex of OC-Ca-P after cropping management. • Cropping management generates more fine colloidal clays, oxides and associated P. Soil phosphorus (P) is usually hardly present in soluble forms as most of it is strongly bound to minerals. Therefore, transformations of soil colloidal P can play a key role in enhancing soil fertility. Here, we examined water extracts from a chronosequence of soils with 2000 years of paddy rice and 700 years of non-paddy cropping, and fractionated them into <1200 nm, <450 nm, and <3 kDa size fractions using sequential ultrafiltration. Asymmetric-flow field-flow fractionation (AF4) coupled to an organic carbon detector (OCD) and inductively coupled plasma mass spectrometry (ICP-MS) was used for organic carbon and elemental screening in the 3 kDa-450 nm colloidal size range. We found that P increased in all colloidal fractions both with long-term paddy rice and non-paddy managements. Furthermore, P in soil colloids of 3 kDa-450 nm size, as fractionated by AF4, peaked in three size ranges: nanoparticles (NP, ∼3 kDa-20 nm), fine colloids (FC, ∼20–225 nm), and medium-sized colloids (MC, ∼225–450 nm). The NP fraction originally contained mainly P and Ca for tidal wetland (TW), but turned into a complex of OC-Ca-P colloidal forms immediately after onset of arable management, with little temporal change thereafter. Fine colloidal P (FC-P) was detected only after > 50 years for the non-paddy and > 100 years under paddy management, associated with oxides, organic matter and fine clay fragments in the paddy soils and additionally Ca-bound P in the non-paddy ones, respectively. Increasing portions of medium-sized colloidal P appeared after 50 years paddy and non-paddy management, though not exceeding the concentrations of FC-P and with little changes during prolonged arable land use. Overall, after initial formation of FC-P during the first 50–100 years of management, longer term dynamics of colloidal P in these arable soils was fairly independent of paddy or non-paddy managements, but remained closely coupled to the increased presence or generation of fine colloidal clays, oxides and the associated P, thus potentially linking P fertility largely to the dynamics of fine colloids. [ABSTRACT FROM AUTHOR]

Size distribution of humic substances is a central issue when one considers stability against aggregation and transportation of HS with bound metal ions. In the present study, the effects of low molecular weight weak organic acids, which are commonly used as pH-buffering agents, on the hydrodynamic size of purified Aldrich humic acid (PAHA) at pH 7 were investigated by means of flow field-flow fractionation (Fl-FFF). The modal size of the PAHA fractogram with 3-morpholino propane sulfonic acid (MOPS) was similar to that obtained without organic acids, whereas that with tris(hydroxymethyl)aminomethane (Tris) increased and that with 2-( N -morpholino)ethanesulfonic acid (MES) decreased relative to that without organic acids. Differences in the charges of the weak organic acids were likely the cause of this difference in the fractograms. After selecting MOPS as an appropriate pH-buffering agent, the size-dependency of metal cation binding (UO 2 2+ and Eu 3+ ) by PAHA was studied at low loading conditions (10 −6 –10 −8 M). The binding of the metal ions was not homogeneous with respect to the size of the PAHA particles; larger particles had greater affinity for both UO 2 2+ and Eu 3+ . The modal size for the hydrodynamic diameter of PAHA was 2 nm, whereas the modal size for the bound metal ions per unit mass of PAHA was 5 nm. This contrast indicates that PAHA particles with 5 nm sizes strongly bind UO 2 2+ and Eu 3+ in terms of their affinity and the number of relevant sites. Size variation of PAHA induced by the addition of multivalent cations was insignificant in the present study. [ABSTRACT FROM AUTHOR]

Hyphenation of asymmetric flow field-flow fractionation (AF4) to an on-line elemental detection (inductively coupled plasma-mass spectrometry, ICP-MS) is proposed as a powerful diagnostic tool for quantum dots bioconjugation studies. In particular, conjugation effectiveness between a “model” monoclonal IgG antibody (Ab) and CdSe/ZnS core-shell Quantum Dots (QDs), surface-coated with an amphiphilic polymer, has been monitored here by such hybrid AF4-ICP-MS technique. Experimental conditions have been optimized searching for a proper separation between the sought bioconjugates from the eventual free reagents excesses employed during the bioconjugation (QDs and antibodies). Composition and pH of the carrier have been found to be critical parameters to ensure an efficient separation while ensuring high species recovery from the AF4 channel. An ICP-MS equipped with a triple quadropole was selected as elemental detector to enable sensitive and reliable simultaneous quantification of the elemental constituents, including sulfur, of the nanoparticulated species and the antibody. The hyphenated technique used provided nanoparticle size-based separation, elemental detection, and composition analysis capabilities that turned out to be instrumental in order to investigate in depth the Ab-QDs bioconjugation process. Moreover, the analytical strategy here proposed allowed us not only to clearly identify the bioconjugation reaction products but also to quantify nanoparticle:antibodies bioconjugation efficiency. This is a key issue in future development of analytical and bioanalytical photoluminescent QDs applications. [ABSTRACT FROM AUTHOR]

Humic substances (HS) play key role in toxic metal binding and protecting aquatic microorganisms from metal-induced stress. Any environmental changes that could alter HS concentration and reactivity can be expected to modify metal complexation and thus affect metal speciation and bioavailability to microalgae. The present study explores the influence of increased solar irradiance on the chemical structures and molecular weight of Elliott soil humic acid (EHA) and the associated consequences for Cd(II), Cu(II) and Pb(II) complexation and intracellular metal content in microalga. The results demonstrate that high radiance doses induce an oxidation of EHA with a formation of low molecular weight acids, an increase of –OH and –COOH group abundance, and a drop in EHA hydrodynamic size and molecular weight. The photo-induced structural changes are accompanied with a release of metal from M–EHA complexes and narrowing their size distribution, which in turn results in an increase of the intracellular Cd, Cu and Pb contents in microalga Chlamydomonas reinhardtii in agreement with the measured free metal ions concentrations. [ABSTRACT FROM AUTHOR]

Colloidal retention characteristics, recovery and size distribution of model macromolecules and natural dissolved organic matter (DOM) were systematically examined using an asymmetrical flow field-flow fractionation (AFlFFF) system under various membrane size cutoffs and carrier solutions. Polystyrene sulfonate (PSS) standards with known molecular weights (MW) were used to determine their permeation and recovery rates by membranes with different nominal MW cutoffs (NMWCO) within the AFlFFF system. Based on a ≥90% recovery rate for PSS standards by the AFlFFF system, the actual NMWCOs were determined to be 1.9 kDa for the 0.3 kDa membrane, 2.7 kDa for the 1 kDa membrane, and 33 kDa for the 10 kDa membrane, respectively. After membrane calibration, natural DOM samples were analyzed with the AFlFFF system to determine their colloidal size distribution and the influence from membrane NMWCOs and carrier solutions. Size partitioning of DOM samples showed a predominant colloidal size fraction in the <5 nm or <10 kDa size range, consistent with the size characteristics of humic substances as the main terrestrial DOM component. Recovery of DOM by the AFlFFF system, as determined by UV-absorbance at 254 nm, decreased significantly with increasing membrane NMWCO, from 45% by the 0.3 kDa membrane to 2–3% by the 10 kDa membrane. Since natural DOM is mostly composed of lower MW substances (<10 kDa) and the actual membrane cutoffs are normally larger than their manufacturer ratings, a 0.3 kDa membrane (with an actual NMWCO of 1.9 kDa) is highly recommended for colloidal size characterization of natural DOM. Among the three carrier solutions, borate buffer seemed to provide the highest recovery and optimal separation of DOM. Rigorous calibration with macromolecular standards and optimization of system conditions are a prerequisite for quantifying colloidal size distribution using the flow field-flow fractionation technique. In addition, the coupling of AFlFFF with fluorescence EEMs could provide new insights into DOM heterogeneity in different colloidal size fractions. [ABSTRACT FROM AUTHOR]

Asymmetrical flow field-flow fractionation (afFFF) was used to investigate the properties of a plant-produced anthrax toxin protective antigen, pp-PA83. The afFFF fractogram consisted of two main peaks with molar masses similar to the molecular mass of pp-PA83 monomer. afFFF carrier solutions strongly influenced the ratio and the intensity of the two main peaks. These differences indicate that conformation changes in the pp-PA83 molecule occurred during the afFFF analysis. Similar fractograms were obtained for different pp-PA83 formulations when the afFFF carrier solution and the protein formulation were the same (or very similar). The data show that in specific cases, afFFF could be used to study protein conformation and document the importance of studying the influence of the carrier solution on afFFF. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 104:612-617, 2015 [ABSTRACT FROM AUTHOR]

This study describes the effect of ionic strength on the molecular structure of hyaluronic acid (HA) in an aqueous solution using flow field-flow fractionation and multiangle light scattering (FlFFF-MALS). Sodium salts of HA (NaHA) raw materials (∼2 × 10 Da) dispersed in different concentrations of NaCl prepared by repeated dilution/ultrafiltration procedures were examined in order to study conformational changes in terms of the relationship between the radius of gyration and molecular weight (MW) and molecular weight distribution (MWD) of NaHA in solution. This was achieved by varying the ionic strength of the carrier solution used in a frit-inlet asymmetrical FlFFF (FIAF4) channel. Experiments showed that the average MW of NaHA increased as the ionic strength of the NaHA solution decreased due to enhanced entanglement or aggregation of HA molecules. Relatively large molecules (greater than ∼5 MDa) did not show a large increase in RMS radius value as the NaCl concentration decreased. Conversely, smaller species showed larger changes, suggesting molecular expansion at lower ionic strengths. When the ionic strength of the FlFFF carrier solution was decreased, the HA species in a salt-rich solution (0.2 M NaCl) underwent rapid molecular aggregation during FlFFF separation. However, when salt-depleted HA samples ( I = 4.66∼0.38 mM) were analyzed with FFF carrier solutions of a high ionic strength, the changes in both molecular structure and size were somewhat reversible, although there was a delay in correction of the molecular structure. [ABSTRACT FROM AUTHOR]

Flow field-flow fractionation (FlFFF) is a liquid-phase separation technique that can separate macromolecules and particles on the basis of differences in their diffusion coefficient, and therefore their size. The membrane in the FlFFF channel is one of the most important elements involved in accomplishing the separation. This mini-review focuses on requirements and important points related to membranes used in FlFFF channels. Unless it is selected properly, the membrane can be a weak point of the technique due to risks such as sample loss because of the interactions between the sample and the membrane. © 2014 Society of Chemical Industry [ABSTRACT FROM AUTHOR]