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Myeong Hee Moon - One of the best experts on this subject based on the ideXlab platform.

  • evaluation of exosome separation from human serum by frit inlet asymmetrical flow field flow fractionation and Multiangle Light Scattering
    Analytica Chimica Acta, 2020
    Co-Authors: Young Beom Kim, Joon Seon Yang, Gwang Bin Lee, Myeong Hee Moon
    Abstract:

    Abstract Exosomes are extracellular vesicles that mediate intercellular communication, immune response, and tumour metastasis. However, exosome isolation from the blood is complicated because their size and density are similar to those of blood lipoproteins. Here, we employed field programming frit-inlet asymmetrical flow field-flow fractionation (FIAF4) coupled with Multiangle Light Scattering (MALS) for the effective separation of exosomes from free unbound proteins and lipoproteins present in serum samples using different pre-treatment methods, namely, a commercial exosome isolation kit, ultracentrifugation (UC), and a simple centrifugation followed by ultrafiltration (UF). Sizes of the eluted exosomes, as calculated by MALS signals, approximated well with the results of batch dynamic Light Scattering of the collected fractions and with the sizes of polystyrene particles. Exosome separation from lipoproteins was validated by western blotting with several markers of exosomes and lipoproteins, followed by proteomic analysis using nanoflow ultrahigh-performance liquid chromatography-electrospray ionisation-tandem mass spectrometry. UC requires relatively large amounts of serum samples (at least 2 mL) but is more efficient at removing lipoproteins. The UF method with a centrifugal concentrator (300 kDa) was found to be more effective in retrieving exosomes with low serum volumes (50 μL). Altogether, this study demonstrates the application of field programming FIAF4 for the isolation/purification of exosomes from proteins and lipoproteins in the serum.

  • Ionic strength effect on molecular structure of hyaluronic acid investigated by flow field-flow fractionation and Multiangle Light Scattering
    Analytical and Bioanalytical Chemistry, 2015
    Co-Authors: Bitnara Kim, Euijin Hwang, Sohee Woo, Young-soo Park, Myeong Hee Moon
    Abstract:

    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 × 10^6 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.

  • characterization of sodium hyaluronate blends using frit inlet asymmetrical flow field flow fractionation and Multiangle Light Scattering
    Analytical and Bioanalytical Chemistry, 2012
    Co-Authors: Muhammad Ali, Il Hwan Cho, Euijin Hwang, Myeong Hee Moon
    Abstract:

    We characterized ultrahigh molecular weight sodium hyaluronate (NaHA) and blended pharmaceutical products containing NaHA using flow field-flow fractionation and Multiangle Light Scattering–differential refractive index (FlFFF-MALS-DRI). NaHA is a water-soluble polysaccharide with a range of molecular weights (MW; 105~108 Da) that is found in body fluids and tissues. NaHA is also used commercially in pharmaceutical and cosmetic applications. We used a frit inlet asymmetrical FlFFF channel to separate aqueous polymers according to their hydrodynamic size, and we used on-line measurements of Light Scattering to obtain the MW distribution (MWD) as well as structural information about NaHA in aqueous solution. In this study, we investigated NaHA and anti-adhesive blend mixtures of NaHA (a commercial NaHA blend mixture containing sodium carboxymethyl cellulose and a new blend with hydroxyethyl starch (HES)) to determine the molecular weight distribution MWD of NaHA and the blend mixtures and to obtain structural information about these compounds in aqueous solution. We also examined the characteristics of NaHA–HES–polylactic-co-glycolic acid film products exposed to gamma radiation for sterilization purposes.

  • flow field flow fractionation and Multiangle Light Scattering for ultrahigh molecular weight sodium hyaluronate characterization
    Journal of Separation Science, 2010
    Co-Authors: Myeong Hee Moon
    Abstract:

    This review describes the utility of flow field-flow fractionation coupled with Multiangle Light Scattering and differential refractive index (FlFFF-MALS-DRI) detection methods for the separation of ultrahigh molecular weight sodium hyaluronate (NaHA) materials and for the characterization of molecular weight distribution as well as structural determination. The sodium salt of hyaluronic acid (HA), NaHA, is a water-soluble polysaccharide with a broad range of molecular weights (10(5) -10(8) ) found in various naturally occurring fluids and tissues. Basic principles of FlFFF-MALS using field programming for the separation of the degraded products of NaHA prepared by treating raw materials with depolymerization or degradation processes such as membrane filtration, enzymatic degradation, ultrasonic degradation, alkaline reaction, irradiation by γ-rays, and thermal treatment for the development of pharmaceutical applications are introduced. Changes in molecular weight distribution and conformation of NaHA materials due to external stimuli are also discussed.

  • depolymerization study of sodium hyaluronate by flow field flow fractionation Multiangle Light Scattering
    Analytical and Bioanalytical Chemistry, 2009
    Co-Authors: Ji Hye Kwon, Euijin Hwang, Il Hwan Cho, Myeong Hee Moon
    Abstract:

    Thermal depolymerization of ultrahigh-molecular-weight (UHMW) sodium hyaluronate (NaHA) was studied systematically by using frit-inlet asymmetrical flow field-flow fractionation/Multiangle Light Scattering/differential refractive index (FI-AFlFFF/MALS/DRI). FI-AFlFFF was utilized for the size separation of NaHA samples which had been thermally degraded for varied treatment times, followed by Light-Scattering detection to determine MW and structural information of degraded NaHA products. Analysis of NaHA products showed time-dependent depolymerization of raw molecules into smaller-MW components, as well as unfolding of compact structures of UHMW NaHA. To determine whether the observed decrease in MW of sodium hyaluronate originated from the chain degradation of UHMW molecules or from dissociation of entangled complex particles that may have been formed by intermolecular association, narrow size fractions (1 × 107–6 × 107 and >6 × 107 MW) of NaHA molecules were collected during FlFFF separation and followed by thermal treatment. Subsequent FI-AFlFFF/MALS analysis of collected fractions after thermal treatment suggested that the ultrahigh-MW region (>107 Da) of NaHA is likely to result from supermolecular structures formed by aggregation of large molecules.

Karlgustav Wahlund - One of the best experts on this subject based on the ideXlab platform.

  • Size separations of starch of different botanical origin studied by asymmetrical-flow field-flow fractionation and Multiangle Light Scattering
    Analytical and Bioanalytical Chemistry, 2011
    Co-Authors: Karlgustav Wahlund, Mats Leeman, Stalin Santacruz
    Abstract:

    Asymmetrical-flow field-flow fractionation combined with Multiangle Light Scattering and refractive index detection has been revealed to be a powerful tool for starch characterization. It is based on size separation according to the hydrodynamic diameter of the starch components. Starch from a wide range of different botanical sources were studied, including normal starch and high-amylose and high-amylopectin starch. The starch was dissolved by heat treatment at elevated pressure in a laboratory autoclave. This gave clear solutions with no granular residues. Amylose retrogradation was prevented by using freshly dissolved samples. Programmed cross flow starting at 1.0 mL min^−1 and decreasing exponentially with a half-life of 4 min was utilised. The starches showed two size populations representing mainly amylose and mainly amylopectin with an overlapping region where amylose and amylopectin were possibly co-eluted. Most of the first population had molar masses below 10^6 g mol^−1, and most of the second size population had molar masses above 10^7 g mol^−1. Large differences were found in the relative amounts of the two populations, the molar mass, and hydrodynamic diameters, depending on the plant source and its varieties.

  • accuracy in Multiangle Light Scattering measurements for molar mass and radius estimations model calculations and experiments
    Analytical Chemistry, 2003
    Co-Authors: Mats Andersson, Bengt Wittgren, Karlgustav Wahlund
    Abstract:

    Multiangle Light Scattering (MALS) is a well-established technique used to determine the size of macromolecules and particles. In this study, different extrapolation procedures used in MALS were investigated with regard to accuracy and robustness in the obtained molar mass and rms radius. Three different mathematical transformations of the Light Scattering function referred to as the Debye, Zimm, and Berry methods for constructing the Debye plot were investigated for two idealized polymer shapes, homogeneous spheres and random coils, with radii from 25 to 250 nm. The effect of the angular interval used for the extrapolation was investigated, as was the robustness of the different transformations toward errors in the measured Light Scattering intensity at low angles. For an rms radius less than 50 nm, the relative error in molar mass due to extrapolation was less than 1% independent of the method used. For larger radii, the error increased and the extrapolation procedure became more critical. For random co...

  • accuracy in Multiangle Light Scattering measurements for molar mass and radius estimations model calculations and experiments
    Analytical Chemistry, 2003
    Co-Authors: Mats Andersson, Bengt Wittgren, Karlgustav Wahlund
    Abstract:

    Multiangle Light Scattering (MALS) is a well-established technique used to determine the size of macromolecules and particles. In this study, different extrapolation procedures used in MALS were investigated with regard to accuracy and robustness in the obtained molar mass and rms radius. Three different mathematical transformations of the Light Scattering function referred to as the Debye, Zimm, and Berry methods for constructing the Debye plot were investigated for two idealized polymer shapes, homogeneous spheres and random coils, with radii from 25 to 250 nm. The effect of the angular interval used for the extrapolation was investigated, as was the robustness of the different transformations toward errors in the measured Light Scattering intensity at low angles. For an rms radius less than 50 nm, the relative error in molar mass due to extrapolation was less than 1% independent of the method used. For larger radii, the error increased and the extrapolation procedure became more critical. For random coil polymers, the Berry method was superior in terms of accuracy and robustness. For spheres, the Debye method was superior. The Zimm method was inferior to the others. The different extrapolation methods were evaluated and compared on experimental data from a size exclusion chromatography-MALS analysis of an ultrahigh molar mass poly(ethylene oxide) (PEO). The PEO data qualitatively verified the calculations and stressed the importance of optimizing the extrapolation procedure after careful evaluation of the experimental data. A discussion of how to detect erroneous data in an experimental Debye plot is given.

  • polysaccharide characterization by flow field flow fractionation Multiangle Light Scattering initial studies of modified starches
    International Journal of Polymer Analysis and Characterization, 2002
    Co-Authors: Bengt Wittgren, Mats Andersson, Karlgustav Wahlund, Cecilia Arfvidsson
    Abstract:

    Chemically modified starches are commonly used for various purposes. Depending on the type of derivatization, a chemical degradation of the original polymeric structure may occur, resulting in a change of molar mass. It is therefore always of interest to know the molar mass and possibly the conformation of the derivative. Four commercially available hydroxypropyl and hydroxyethyl modified starches were examined by asymmetrical flow field-flow fractionation combined with Multiangle laser Light Scattering. The weight-average molar mass and the molar mass distribution were determined, with emphasis put on the rapid analysis and studies of the suitable experimental conditions regarding flow rates so that accurate data were obtained. The molar mass distribution determinations showed good reproducibility, and repeatability and were fast. Efforts to obtain conformational information are described. (Less)

  • ultrahigh molar mass component detected in ethylhydroxyethyl cellulose by asymmetrical flow field flow fractionation coupled to Multiangle Light Scattering
    Analytical Chemistry, 2001
    Co-Authors: Mats Andersson, Bengt Wittgren, Karlgustav Wahlund
    Abstract:

    Asymmetrical flow field-flow fractionation (flow FFF) was connected to Multiangle Light Scattering (MALS) and refractive index (RI) detectors for characterization of the molar mass distribution and molecular radius of a cellulose derivative, ethylhydroxyethyl cellulose (EHEC). Experimental conditions were optimized to allow study of a wide range of molar mass including even ultrahigh molar mass (UHM) components. The weight-average molar mass was 3.1 x 10(5) g x mol(-1) representing a very broad range (of molar mass) from 4.0 x 10(4) to 10(7) g x mol(-1), which corresponds to from <20 to 200 nm rms radius. The Light Scattering signal showed the presence of an UHM component, possibly an aggregate of extreme size, i.e., approximately 10(8) g x mol(-1) with a hydrodynamic diameter of 0.35 microm. Careful choice of the pore size in in-line filters is necessary in order to minimize MALS detector noise without removing the UHM component. Flow FFF-MALS-RI was demonstrated to be uniquely suited to detect the presence of UHM components.

Conrad Coester - One of the best experts on this subject based on the ideXlab platform.

  • asymmetrical flow field flow fractionation and Multiangle Light Scattering for analysis of gelatin nanoparticle drug carrier systems
    Analytical Chemistry, 2004
    Co-Authors: Wolfgang Fraunhofer, Gerhard Winter, Conrad Coester
    Abstract:

    The physicochemical properties of nanosized colloidal drug carrier systems are of great influence on drug efficacy. Consequently, a broad spectrum of analytical techniques is applied for comprehensive drug carrier characterization. It is the primary objective of this paper to present asymmetrical flow field-flow fractionation (AF4), coupled online with Multiangle Light Scattering detection, for the characterization of gelatin nanoparticles. Size and size distribution of drug-loaded and unloaded nanoparticles were determined, and data were correlated with results of state-of-the-art methods, such as scanning electron microscopy and photon correlation spectroscopy. Moreover, the AF4 fractionation of gelatin nanoparticulate carriers from a protein model drug is demonstrated for the first time, proposing a feasible way to assess the amount of loaded drug in situ without sample preparation. This hypothesis was set into practice by monitoring the drug loading of nanoparticles with oligonucleotide payloads. In t...

  • Asymmetrical Flow Field-Flow Fractionation and Multiangle Light Scattering for Analysis of Gelatin Nanoparticle Drug Carrier Systems
    Analytical Chemistry, 2004
    Co-Authors: Wolfgang Fraunhofer, Gerhard Winter, Conrad Coester
    Abstract:

    The physicochemical properties of nanosized colloidal drug carrier systems are of great influence on drug efficacy. Consequently, a broad spectrum of analytical techniques is applied for comprehensive drug carrier characterization. It is the primary objective of this paper to present asymmetrical flow field-flow fractionation (AF4), coupled online with Multiangle Light Scattering detection, for the characterization of gelatin nanoparticles. Size and size distribution of drug-loaded and unloaded nanoparticles were determined, and data were correlated with results of state-of-the-art methods, such as scanning electron microscopy and photon correlation spectroscopy. Moreover, the AF4 fractionation of gelatin nanoparticulate carriers from a protein model drug is demonstrated for the first time, proposing a feasible way to assess the amount of loaded drug in situ without sample preparation. This hypothesis was set into practice by monitoring the drug loading of nanoparticles with oligonucleotide pay-loads. In this realm, various fractions of gelatin bulk material were analyzed via AF4 and size-exclusion high-pressure liquid chromatography. Mass distributions and high-molecular-weight fraction ratios of the gelatin samples varied, depending on the separation method applied. In general, the AF4 method demonstrated the ability to comprehensively characterize polymeric gelatin bulk material as well as drug-loaded and unloaded nanoparticles in terms of size, size distribution, molecular weight, and loading efficiency.

Wolfgang Fraunhofer - One of the best experts on this subject based on the ideXlab platform.

  • asymmetrical flow field flow fractionation and Multiangle Light Scattering for analysis of gelatin nanoparticle drug carrier systems
    Analytical Chemistry, 2004
    Co-Authors: Wolfgang Fraunhofer, Gerhard Winter, Conrad Coester
    Abstract:

    The physicochemical properties of nanosized colloidal drug carrier systems are of great influence on drug efficacy. Consequently, a broad spectrum of analytical techniques is applied for comprehensive drug carrier characterization. It is the primary objective of this paper to present asymmetrical flow field-flow fractionation (AF4), coupled online with Multiangle Light Scattering detection, for the characterization of gelatin nanoparticles. Size and size distribution of drug-loaded and unloaded nanoparticles were determined, and data were correlated with results of state-of-the-art methods, such as scanning electron microscopy and photon correlation spectroscopy. Moreover, the AF4 fractionation of gelatin nanoparticulate carriers from a protein model drug is demonstrated for the first time, proposing a feasible way to assess the amount of loaded drug in situ without sample preparation. This hypothesis was set into practice by monitoring the drug loading of nanoparticles with oligonucleotide payloads. In t...

  • Asymmetrical Flow Field-Flow Fractionation and Multiangle Light Scattering for Analysis of Gelatin Nanoparticle Drug Carrier Systems
    Analytical Chemistry, 2004
    Co-Authors: Wolfgang Fraunhofer, Gerhard Winter, Conrad Coester
    Abstract:

    The physicochemical properties of nanosized colloidal drug carrier systems are of great influence on drug efficacy. Consequently, a broad spectrum of analytical techniques is applied for comprehensive drug carrier characterization. It is the primary objective of this paper to present asymmetrical flow field-flow fractionation (AF4), coupled online with Multiangle Light Scattering detection, for the characterization of gelatin nanoparticles. Size and size distribution of drug-loaded and unloaded nanoparticles were determined, and data were correlated with results of state-of-the-art methods, such as scanning electron microscopy and photon correlation spectroscopy. Moreover, the AF4 fractionation of gelatin nanoparticulate carriers from a protein model drug is demonstrated for the first time, proposing a feasible way to assess the amount of loaded drug in situ without sample preparation. This hypothesis was set into practice by monitoring the drug loading of nanoparticles with oligonucleotide pay-loads. In this realm, various fractions of gelatin bulk material were analyzed via AF4 and size-exclusion high-pressure liquid chromatography. Mass distributions and high-molecular-weight fraction ratios of the gelatin samples varied, depending on the separation method applied. In general, the AF4 method demonstrated the ability to comprehensively characterize polymeric gelatin bulk material as well as drug-loaded and unloaded nanoparticles in terms of size, size distribution, molecular weight, and loading efficiency.

Bengt Wittgren - One of the best experts on this subject based on the ideXlab platform.

  • accuracy in Multiangle Light Scattering measurements for molar mass and radius estimations model calculations and experiments
    Analytical Chemistry, 2003
    Co-Authors: Mats Andersson, Bengt Wittgren, Karlgustav Wahlund
    Abstract:

    Multiangle Light Scattering (MALS) is a well-established technique used to determine the size of macromolecules and particles. In this study, different extrapolation procedures used in MALS were investigated with regard to accuracy and robustness in the obtained molar mass and rms radius. Three different mathematical transformations of the Light Scattering function referred to as the Debye, Zimm, and Berry methods for constructing the Debye plot were investigated for two idealized polymer shapes, homogeneous spheres and random coils, with radii from 25 to 250 nm. The effect of the angular interval used for the extrapolation was investigated, as was the robustness of the different transformations toward errors in the measured Light Scattering intensity at low angles. For an rms radius less than 50 nm, the relative error in molar mass due to extrapolation was less than 1% independent of the method used. For larger radii, the error increased and the extrapolation procedure became more critical. For random co...

  • accuracy in Multiangle Light Scattering measurements for molar mass and radius estimations model calculations and experiments
    Analytical Chemistry, 2003
    Co-Authors: Mats Andersson, Bengt Wittgren, Karlgustav Wahlund
    Abstract:

    Multiangle Light Scattering (MALS) is a well-established technique used to determine the size of macromolecules and particles. In this study, different extrapolation procedures used in MALS were investigated with regard to accuracy and robustness in the obtained molar mass and rms radius. Three different mathematical transformations of the Light Scattering function referred to as the Debye, Zimm, and Berry methods for constructing the Debye plot were investigated for two idealized polymer shapes, homogeneous spheres and random coils, with radii from 25 to 250 nm. The effect of the angular interval used for the extrapolation was investigated, as was the robustness of the different transformations toward errors in the measured Light Scattering intensity at low angles. For an rms radius less than 50 nm, the relative error in molar mass due to extrapolation was less than 1% independent of the method used. For larger radii, the error increased and the extrapolation procedure became more critical. For random coil polymers, the Berry method was superior in terms of accuracy and robustness. For spheres, the Debye method was superior. The Zimm method was inferior to the others. The different extrapolation methods were evaluated and compared on experimental data from a size exclusion chromatography-MALS analysis of an ultrahigh molar mass poly(ethylene oxide) (PEO). The PEO data qualitatively verified the calculations and stressed the importance of optimizing the extrapolation procedure after careful evaluation of the experimental data. A discussion of how to detect erroneous data in an experimental Debye plot is given.

  • polysaccharide characterization by flow field flow fractionation Multiangle Light Scattering initial studies of modified starches
    International Journal of Polymer Analysis and Characterization, 2002
    Co-Authors: Bengt Wittgren, Mats Andersson, Karlgustav Wahlund, Cecilia Arfvidsson
    Abstract:

    Chemically modified starches are commonly used for various purposes. Depending on the type of derivatization, a chemical degradation of the original polymeric structure may occur, resulting in a change of molar mass. It is therefore always of interest to know the molar mass and possibly the conformation of the derivative. Four commercially available hydroxypropyl and hydroxyethyl modified starches were examined by asymmetrical flow field-flow fractionation combined with Multiangle laser Light Scattering. The weight-average molar mass and the molar mass distribution were determined, with emphasis put on the rapid analysis and studies of the suitable experimental conditions regarding flow rates so that accurate data were obtained. The molar mass distribution determinations showed good reproducibility, and repeatability and were fast. Efforts to obtain conformational information are described. (Less)

  • ultrahigh molar mass component detected in ethylhydroxyethyl cellulose by asymmetrical flow field flow fractionation coupled to Multiangle Light Scattering
    Analytical Chemistry, 2001
    Co-Authors: Mats Andersson, Bengt Wittgren, Karlgustav Wahlund
    Abstract:

    Asymmetrical flow field-flow fractionation (flow FFF) was connected to Multiangle Light Scattering (MALS) and refractive index (RI) detectors for characterization of the molar mass distribution and molecular radius of a cellulose derivative, ethylhydroxyethyl cellulose (EHEC). Experimental conditions were optimized to allow study of a wide range of molar mass including even ultrahigh molar mass (UHM) components. The weight-average molar mass was 3.1 x 10(5) g x mol(-1) representing a very broad range (of molar mass) from 4.0 x 10(4) to 10(7) g x mol(-1), which corresponds to from <20 to 200 nm rms radius. The Light Scattering signal showed the presence of an UHM component, possibly an aggregate of extreme size, i.e., approximately 10(8) g x mol(-1) with a hydrodynamic diameter of 0.35 microm. Careful choice of the pore size in in-line filters is necessary in order to minimize MALS detector noise without removing the UHM component. Flow FFF-MALS-RI was demonstrated to be uniquely suited to detect the presence of UHM components.

  • conformational change and aggregation of κ carrageenan studied by flow field flow fractionation and Multiangle Light Scattering
    Biopolymers, 1998
    Co-Authors: Bengt Wittgren, J Borgstrom, Lennart Piculell, K G Wahlund
    Abstract:

    The relatively novel combination of flow field-flow fractionation (FFF) and Multiangle Light Scattering (MALS) was employed to study a nondegraded κ-carrageenan in different 0.1M salt solutions. The applicability of the technique was tested, and the effects of salt type and salt composition on the molar mass and radius of gyration were studied. A conformational ordering was induced at room temperature by switching the solvent from 0.1M NaCl (coil form) to 0.1M NaI (helix form). An approximate doubling of the average molar mass and an increase in radius of gyration was then observed, in agreement with results obtained previously using size exclusion chromatography–MALS. This increase in size was attributed to conformational ordering and to the formation of double helices. Severe aggregation was observed above 40% CsI in the 0.1M mixed salt solution of CsI and NaI. This was ascribed to the association of helices into large aggregates. For these large associates, having molar masses of several millions, a reversal of the elution order in flow FFF was detected. © 1998 John Wiley & Sons, Inc. Biopoly 45: 85–96 1998