Rapid detection of compositional drift of polydisperse copolymers using thermal field-flow fractionation and multi-angle light scattering

Summary The use of thermal field-flow fractionation (ThFFF) with multi-angle light scattering (MALS) for the rapid detection of compositional heterogeneity in random copolymers is demonstrated. Soret coefficients were directly calculated from the ThFFF retention times while the MALS detector provided the polymer's radius of gyration (R _g) distribution. FromR _g, the diffusion coefficient (D) could be calculated and this allowed, in combination with the Soret coefficient, the calculation of the thermal diffusion coefficient (D _T). It was shown that theD _T distribution can serve as a measure for the chemical composition distribution of random styrene acrylonitrile copolymers. Comparison of ThFFF-MALS results with literature data from ThFFF-hydrodynamic chromatography (HDC) cross-fractionation experiments showed a fair agreement.     

Contribution of flow field flow fractionation with on line static and dynamic light scattering to the study of hydrosoluble polyelectrolyte complexes

Water soluble polyelectrolyte complexes (PECs) formed between polyaspartate (anionic polymer) and poly(trimethylammonium propyl methacrylamide chloride) (cationic polymer) were studied by flow field flow fractionation with on-line coupling multi-angle laser light scattering-quasi elastic light scattering-differential refractive index determination (F4/MALLS/QELS/DRI). The separation technique permits to characterize polydisperse PECs. The molar mass of the polycation (PC) influences the stiffness of the PECs and the proportion between single PECs (i.e. nPA/1PC) and multiple PECs (i.e. nPA/n’PC). High ionic strength with NaCl (>0.1 M) tends to break the multiple PECs while CaCl₂ destroys PECs and leads to the formation of complexes polyaspartate/Ca²⁺. The studied PECs can be used as inhibitors to the calcite formation in the drilling fluids.     

Analysis of liposomes using asymmetrical flow field-flow fractionation: Separation conditions and drug/lipid recovery

Liposomes composed of dipalmitoylphosphatidylcholine and dipalmitoylphosphatidylglycerol were analyzed by asymmetrical flow field-flow fractionation coupled with multi-angle laser light scattering. In addition to evaluation of fractionation conditions (flow conditions, sample mass, carrier liquid), radiolabeled drug-loaded liposomes were used to determine the liposome recovery and a potential loss of incorporated drug during fractionation. Neither sample concentration nor the cross-flow gradient distinctly affected the size results but at very low sample concentration (injected mass 5 μg) the fraction of larger vesicles was underestimated. Imbalance in the osmolality between the inner and outer aqueous phase resulted in liposome swelling after dilution in hypoosmotic carrier liquids. In contrast, liposome shrinking under hyperosmotic conditions was barely visible. The liposomes themselves eluted completely (lipid recoveries were close to 100%) but there was a loss of incorporated drugs during separation with a strong dependence on the octanol-water partition coefficient of the drug. Whereas corticosterone (partition coefficient ～2) was washed out more or less completely (recovery about 2%), loss of temoporfin (partition coefficient ～9) was only minor (recovery about 80%). All fractionations were well repeatable under the experimental conditions applied in the present study.     

Analysis of a polydisperse system using small-angle X-ray scattering

The small-angle X-ray data from a polydisperse solution of sodium silicate have been measured in the concentration range 3.6–169 mg/cm³ using aKratky camera. The following values of the particle parameters were obtained: the average radius of gyration =7.5 nm, the average particle weight =900 000, the average volume =671 nm³, and the average particle surface area =717 nm².From the above parameters and the apparent specific volume, analysed to be 0.422 cm³/g, the water content of the silicate particles was determined to be 3% (by weight).From small-angle X-ray measurements, performed on solutions exposed to a hydrodynamic field, it is indicated that at least the larger particles in the solution have a relatively symmetric shape. Based on this observation it was assumed that the particles in solution are spherical, and particle size distribution functions were calculated using a least-squares program. It was found that the distribution cannot be described by a simple function, such as aGaussian function; instead, the distribution follows a histogram with three local maxima.Dedicated to Prof. Dr. Dr.Otto Kratky, Graz, on the occasion of his 80th birthday.     

Quantification of uncertainties involved in the conformational study of polymers by light scattering

Gel permeation chromatography (GPC) combined with a multi-angle light scattering (MALS) is a very powerful characterization technique because it provides both absolute molecular weight (M_w) and the radius of gyration (R_g) throughout the separated slices obtained by GPC. This combination of M_w and R_g, can be used to obtain information about the conformation of polymer chains in solutions and the branching of molecules. Due to the interesting properties obtained for polymers, it is essential to quantify the effect of different error sources in light scattering measurements as well as in the data treatment that highly affect the accuracy of obtained molar mass and radius of gyration. Usually, the results obtained for M_w and R_g in this analysis are dispersed for determined ranges of retention time and to have both reliable R_g and M_w for calculation, only high confidence data points have to be chosen. This range is arbitrarily chosen by the user for the data observation.In this work a new method of calculation to obtain R_g and M_w by means of GPC–MALS technique has been developed. As a first point, a data analysis procedure was set in order to describe both R_g and M_w vs. retention time and to determine the range where experimental data are confident. Several aspects in the data analysis have been studied. The polynomial fit function, the influence of the concentration of the sample, the reproducibility of the experiments and the conformational scaling law have been investigated by statistic technique in order to quantify the uncertainties involved.     

Asymmetric flow field flow fractionation of aqueous C60 nanoparticles with size determination by dynamic light scattering and quantification by liquid chromatography atmospheric pressure photo-ionization mass spectrometry

A size separation method was developed for aqueous C₆₀ fullerene aggregates (aqu/C₆₀) using asymmetric flow field flow fractionation (AF4) coupled to a dynamic light scattering detector in flow through mode. Surfactants, which are commonly used in AF4, were avoided as they may alter suspension characteristics. Aqu/C₆₀ aggregates generated by sonication in deionized water ranged in size from 80 to 260 nm in hydrodynamic diameter (D_h) as determined by DLS in flow through mode, which was corroborated by analysis of fractions by DLS in batch mode and by TEM. The mass of C₆₀ in each fraction was determined by LC–APPI–MS. Only 5.2 ± 6.7% of the total aqu/C₆₀ mass had D_h less than 80 nm, while 58 ± 32% of the total aqu/C₆₀ mass had D_h between 80 and 150 nm and 14 ± 9.2% of the total aqu/C₆₀ were between 150 and 260 nm in D_h. With the optimal fractionation parameters, 77 ± 5.8% of the aqu/C₆₀ mass eluted from the AF4 channel, indicating deposition on the AF4 membrane had occurred during fractionation; use of alternative membranes did not reduce deposition. Channel flow splitting increased detector response although channel split ratios greater than 80% of the channel flow led to decreased detector response. This is the first report on the use of AF4 for fractionating a colloidal suspension of aqu/C₆₀.     

Asymmetrical flow field-flow fractionation with multi-angle light scattering detection for the analysis of structured nanoparticles

Synthesis and applications of new functional nanoparticles are topics of increasing interest in many fields of nanotechnology. Chemical modifications of inorganic nanoparticles are often necessary to improve their features as spectroscopic tracers or chemical sensors, and to increase water solubility and biocompatibility for applications in nano-biotechnology. Analysis and characterization of structured nanoparticles are then key steps for their synthesis optimization and final quality control. Many properties of structured nanoparticles are size-dependent. Particle size distribution analysis then provides fundamental analytical information. Asymmetrical flow field-flow fractionation (AF4) with multi-angle light scattering (MALS) detection is able to size-separate and to characterize nanosized analytes in dispersion. In this work we focus on the central role of AF4-MALS to analyze and characterize different types of structured nanoparticles that are finding increasing applications in nano-biotechnology and nanomedicine: polymer-coated gold nanoparticles, fluorescent silica nanoparticles, and quantum dots. AF4 not only size-fractionated these nanoparticles and measured their hydrodynamic radius (r_h) distribution but it also separated them from the unbound, relatively low-M_r components of the nanoparticle structures which were still present in the sample solution. On-line MALS detection on real-time gave the gyration radius (r_g) distribution of the fractionated nanoparticles. Additional information on nanoparticle morphology was then obtained from the r_h/r_g index. Stability of the nanoparticle dispersions was finally investigated. Aggregation of the fluorescent silica nanoparticles was found to depend on the concentration at which they were dispersed. Partial release of the polymeric coating from water-soluble QDs was found when shear stress was induced by increasing flowrates during fractionation.     

Depolymerization study of sodium hyaluronate by flow field-flow fractionation/multiangle light scattering

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.     

Contribution to polymer nanoparticles analysis by laser light scattering

The applications of laser light scattering (LLS) to polymer physics and colloid science have been extensive and noteworthy, especially in particle-size analysis. The study presents an example of LLS application to the characterization of interpolymer complexation of poly(aspartic acid) with a vinylic polymer, by particle size and zeta potential evaluation. The LLS technique is complemented by oscillatory rheological data. The complexes have a potential biomedical application by bioactive substances attachment.     

Enhanced resonance light scattering properties of gold nanoparticles due to cooperative binding

The interaction of 11-mercaptoundecanoic acid capped gold nanoparticles (MUA-GNPs) with europium ions and aminoacids has been studied by UV-Vis spectrophotometry, fluorescence, confocal fluorescence microscopy, resonance light scattering and TEM. Results demonstrated that hyper-Rayleigh scattering emission occurs upon the addition of lysine to the MUA-GNPs–Eu(III) system, thus providing an inherently sensitive method for lysine determination. The effects of geometrical factors of the gold nanoparticles (aspect ratio, particle size, cluster formation) and the surrounding medium (pH) on this behavior are discussed. The cooperative binding interactions of Eu³⁺ and lysine with gold nanoparticles permitted the discrimination of lysine from other amino acids. The probable mechanism for the spectral changes and the enhanced resonance light scattering observed is outlined. Figure Gold nanoparticle resonance light scattering plasmon enhancement through cooperative binding with europium and lysine     

Using two-dimensional time resolved light scattering to study the cure reaction induced phase separation process of epoxy-amine-polyethersulfone blend with secondary phase separation

<正>The generalized two-dimensional correlation analysis based on time-resolved light scattering patterns(2D TRLS) has been employed to study the phase separation process of an epoxy-amine-polyethersulfone blend in which the secondary phase separation takes place.The results of the 2D TRLS provided more detailed information that was not readily observed in the 1D TRLS patterns.(i) During the first process of phase separation,the sequential order of coarsening in size of the domains among the larger and smaller ones has been reversed between the diffusion regime and the hydrodynamic regime. (ii) The change of the larger domains in size,due to the hydrodynamic flow in the late stage of the first phase separation process,keeps on taking place earlier than that of the new domains appeared in the secondary phase separation process. (iii) During the secondary phase separation process the size growth of the smaller domains takes place earlier than that of the larger ones,probably due to the assumption that the coarsening mode could decrease the interface tension more quickly.     

银的流动注射共振光散射法测定

利用自制的流通池,研究了流动注射分析与共振光散射光谱法联用的测定技术;探讨了Ag的流动注射和共振光散射光谱测定的工作条件;Ag的线性范围为0～100 μg/mL,检出限为0.136 μg/mL,测定频率为43次/h;实验结果表明本文所建立的方法快速、所需的化学试剂少.     

Glucose detection at attomole levels using dynamic light scattering and gold nanoparticles

Glucose is directly related to brain activity and to diabetes.Therefore,developing a rapid and sensitive method for glucose detection is essential.Here,label-free glucose detection at attomole levels was realized by detecting the average diameter change of gold nanoparticles(AuNPs)utilizing dynamic light scattering(DLS).Single-strand DNA(ssDNA)adsorbed into the AuNPs’surfaces and prevented them from aggregating in solution that contained NaCl.However,ssDNA cleaved onto ssDNA fragments upon addition of glucose,and these fragments could not adsorb onto the AuNPs’surfaces.Therefore,in high-salt solution,AuNPs would aggregate and their average diameter would increase.Based on monitoring the average diameter of AuNPs with DLS,glucose could be detected in the range from 15 pmol/L to 2.0 nmol/L,with a detection limit of 8.3 pmol/L.Satisfactory results were also obtained when the proposed method was applied in human serum glucose detection.     

Laser light scattering study of the degradation of poly(sebacic anhydride) nanoparticles

Poly(sebacic anhydride) (PSA) is biocompatible and degradable in basic media. We micronized this water‐insoluble polymer into stable polymeric nanoparticles via a microphase inversion. Such PSA nanoparticles degraded much faster than bulk PSA. The influence of the surfactant, temperature, and pH on the degradation of the PSA nanoparticles was investigated by a combination of static and dynamic laser light scattering. Under each condition, the degradation rate was nearly constant up to a 75% weight loss; that is, the degradation was close to zero‐order. The degradation rate increased with the pH and temperature. Biomedical applications of such PSA nanoparticles are suggested. © 2001 John Wiley & Sons, Inc. J Polym Sci B: Polym Phys 39: 703–708, 2001     

A flow injection sampling resonance light scattering system for total protein determination in human serum

A novel flow injection method with resonance light scattering detection was developed for the determination of total protein concentrations. This method is based on the enhancement of RLS signals from Methyl Blue (MB) by protein. The enhanced RLS intensities at 333 nm, in a pH 4.1 acidic aqueous solution, were proportional to the protein concentration over the range 2.0-37.3 and 1.0-36.0 microg ml-1 for human serum albumin (HSA) and bovine serum albumin (BSA), respectively. The corresponding limits of detection (3sigma) of 45 ng ml-1 for HSA and 80 ng ml-1 for BSA were attained. The method was successfully applied to the quantification of total proteins in human serum samples, the maximum relative error is less than 1% and the recovery is between 98% and 102%. The sample throughput was 60 h-1.     

Determination of proteins by flow injection analysis coupled with the Rayleigh light scattering technique

A novel flow injection analysis (FIA) method with Rayleigh light scattering (RLS) detection was developed for the determination of protein concentrations. This method is based on the weak intensity of RLS of p-nitrohenzene-azo-3,6 disulfo-1-amino-8-naphthol-7-azo-benzene disodium salt (Amide Black-10B) which can be enhanced by addition of protein in weakly acidic solution. It has proved that the application of this method to quantify the proteins by using human serum albumin was available in real samples. In addition, this method is very sensitive (the determination limits are 0.11 μg/mL for human serum albumen (HSA) and 0.85 μg/mL for bovine serum albumen, BSA), simple, rapid and tolerance of most interfering substances. The FIA-RLS method was more stabile than the general RLS method and the average R.S.D. value of FIA-RLS less than general RLS. The effects of different interfering substances will be also examined. The amount of proteins in human serum sample was determined and the maximum relative error was no more than 3.00% as well as the recovery was between 94.9 and 105.9%.     

Electrochemical response and separation in cyclic electric field-flow fractionation

Electric field-flow fractionation (EFFF) is a separation technique that couples a lateral electric field with axial Poiseuille flow to separate particles on the basis of size and/or mobility. In unidirectional EFFF, the field rapidly decreases in time due to charging of the double layer. The field strength could be increased by performing EFFF with cyclic electric fields. In cyclic electric field-flow fractionation (CEFFF), a periodic voltage, which can be either sinusoidal or square-wave, is applied in the lateral direction. In this paper, we measure the electrochemical response of CEFFF, i.e., the current-time response for a given time-dependent voltage and then utilize this electrochemical response in a transport model to predict separation. The CEFFF device studied here comprises two gold-coated glass plates separated by a spacer. The transient current profiles are measured for a step change and cyclic square-shaped voltage. The current profile is compared with the equivalent circuit model, and is fitted to a sum of two decaying exponentials. The dependence of the electrochemical response on voltage, frequency, channel thickness, and salt concentration is studied. Next, the electrochemical data are utilized in the convection-diffusion equation to develop a model for separation by CEFFF. The equations are solved by using a combination of analytical and numerical techniques to determine the mean velocity and the dispersion coefficient of molecules, and to determine the effect of various parameters on the separation efficiency of the EFFF device. Also, the model predictions are compared with experimental data available in the literature.     

Comments on the separation efficiency of asymmetrical flow field-flow fractionation in channels of constant channel and crossflow velocities leading to constant separation efficiency

It is shown theoretically that a claim in the literature about the overall separation efficiency of asymmetrical flow FFF channels being improved by geometries that permit a uniform channel flow velocity throughout the channel length is untrue.     

Determination of gamma-globulin at nanogram levels by its enhancement effect on the resonance light scattering of functionalized HgS nanoparticles

A novel assay of γ-globulin (γ-IgG) with a sensitivity at the nanogram level is proposed based on the measurement of enhanced resonance light-scattering (RLS) signals resulting from the interaction of functionalized nano-HgS with γ-globulin. At pH 5.03, the RLS signals of functionalized nano-HgS were greatly enhanced by γ-globulin in the region of 200-700 nm characterized by the peak around 362 nm. Linear relationship can be established between the enhanced RLS intensity and γ-globulin concentration in the range of 10-140 ng ml⁻¹. The limit of detection is 2.71 ng ml⁻¹. Based on this, a new direct quantitative determination method for γ-globulin in blood serum samples without separation of human serum albumin (HSA) is established. The contents of γ-IgG in blood serum samples were determined with recovery of 95.7-102.5% and R.S.D. of 1.6-2.4%. This method is proved to be very sensitive, rapid, simple and tolerance of most interfering substances.     

Analysis of silica nanoparticles by capillary electrophoresis coupled to an evaporative light scattering detector

A simple and rapid methodology has been developed to identify and separate silica nanoparticles (SiO₂NPs) of different sizes in aqueous solution by capillary zone electrophoresis coupled to an evaporative light scattering detector (CE-ELSD). SiO₂NPs were separated using 3 mM ammonium acetate buffer, containing 1% methanol at pH 6.9. SiO₂NPs of 20, 50 and 100 nm were successfully separated under the optimum experimental conditions. CE coupled to ELSD has been proven to be an effective separation technique to determine particles with such small sizes, although the peaks are very close to each other, and it is a promising technique that may allow the separation of other types of nanoparticles. Confirmation by TEM and quantification of the SiO₂ content was also carried out by inductively coupled plasma-mass spectrometry (ICP-MS). The new method was applied to the analysis of real samples, in order to assess its ability to avoid matrix effects in the determination of SiO₂NPs in these kinds of samples.