Bedřich Porsch

Epoxy- and diol-modified silica: Optimization of surface bonding reaction

Abstract The 3-glycidyloxypropyltrimethoxysilane-silica bonding reaction was investigated. The carbon and bonded epoxide content after the bonding reaction and

Size-exclusion chromatography and dynamic light scattering of dextrans in water: Explanation of ion-exclusion behaviour

Abstract The size-exclusion chromatographic (SEC) and ion-exclusion chromatographic (IEC) behaviour of dextrans in water was studied in combination with dynamic light scattering (DLS) of their fractions obtained from micropreparative SEC in water. Dextrans do not aggregate in dilute aqueous solution, but very low dust particle contamination, of particle size 110–170 nm, was revealed from DLS measurements after its separation and its concentration enhancement by SEC. The ion exclusion of charged dextran molecules is the only cause of two excluded peaks observed in the SEC of dextrans in pure water. Model dextran oxidation experiments were performed and mono- and dicarboxydextran were identified as forming these ion-excluded peaks. An IEC-SEC experiment is capable of detecting the presence of polysaccharide chains bearing one or two carboxyl groups only, and of resolving them from each other and from the unchanged chains of molecular masses up to 106 if pure water is used as the mobile phase. The presence of such low charge density macromolecules appears to be a general feature of polysaccharides.

Some specific problems in the practice of preparative high-performance liquid chromatography

Abstract The practice of preparative high-performance liquid chromatography (PLC) is reviewed. Special attention is paid to problems with the use of this method in research and development which are insignificant or unfamiliar on an analytical scale. PLC column concepts, stability and related packing procedures are discussed. Guidelines to column size selection and optimum use are presented. The paramount importance of high resolution for successful PLC separation is stressed and the effect of friction heat generated by viscous flow on the column performance is described. The significance of sufficient sample solubility in the mobile phase is discussed. Possible deleterious effects of the use of strong solvents with viscosities different from that of the mobile phase are considered. The packing solubility is shown to influence product purity; various product isolation procedures are discussed and the use of solid-phase extraction is recommended.

Molar mass distribution of hydroxypropyl cellulose by size exclusion chromatography with dual light scattering and refractometric detection

Physico-chemical properties of cellulose derivatives are of considerable interest in many technical applications, for example, in the food and drug industry. Efficient and careful characterisation of these properties is thus highly desirable. In this study, two different size exclusion chromatography (SEC) systems, connected on-line either to a low-angle laser light scattering detector (LALLS) or to a multi-angle laser light scattering detector (MALLS), are employed for size characterisation of three batches of hydroxypropyl cellulose (HPC) from different manufacturers. All three samples turned out to have a weight average molar mass around 100,000 g/mol, but considerable differences concerning conformational properties were found. Two of the samples contained compact components, presumably aggregates of HPC, which were clearly detected by both SEC-systems. The third sample, obtained from another manufacturer, did not show any indication of aggregation. Both SEC-MALLS and SEC-LALLS are proven to be efficient techniques for characterisation of complex polysaccharides like HPC containing mixtures of solvated polymer chains, as well as micelle-like aggregates.

Ion-exclusion controlled size-exclusion chromatography of methacrylic acid-methyl methacrylate copolymers.

Controlled ion-exclusion allows compensation of hydrophobic adsorption in size-exclusion chromatography of negatively charged methacrylic acid-methyl methacrylate (Eudragit) polymers using methanol as a mobile phase. Properly selected low-ionic-strength conditions below 5 mM LiCl provide correct separation in the size-exclusion mode. Possible disturbing effects, mainly related to light scattering, under low-salt conditions are discussed and shown to be negligible if on-line concentration-light scattering detection is used. The absence of these disturbances is checked by a comparison of experiments performed in methanol containing 1.25 mM and 2.5 mM LiCl. Molecular mass averages and distributions identical within the experimental error are obtained.

Effect of alkalinization on carboxylated latices prepared with various amount of a non-ionogenic hydrophilic comonomer 2-hydroxyethyl methacrylate

Abstract Six latices of carboxylated styrene/butyl acrylate copolymers with 0–25% of 2-hydroxyethyl methacrylate (HEMA) were prepared by semicontinuous non-seeded emulsion polymerization and investigated by photon correlation spectroscopy and viscometry. Although HEMA is non-ionogenic, it profoundly affects the latex behavior after alkalinization: both methods revealed that HEMA promotes an increase in hydrodynamic dimensions of dispersed particles. The increase in viscosity due to incorporation of 10% of HEMA into the copolymers can make more than one order of magnitude at high volume fractions.

Batch and Size-Exclusion Chromatographic Characterization of Ultra-high Molar Mass Sodium Hyaluronate Containing Low Amounts of Strongly Scattering Impurities by Dual Low Angle Light Scattering/Refractometric Detection

Abstract It is shown that very pure sodium hyaluronate (HA) contains small amounts of strongly scattering impurities not detectable on mass scale by refractometric detection, but clearly detectable using low angle light scattering detection during a static bulk light scattering experiment. Size filtration of its solutions does not remove these impurities, only reduces their amount depending on filter porosity. Complete removal of these particle impurities, independent of filter porosity, is achieved by hydrophobic adsorption on hydrophobic filter membranes. Using 0.1 M NaCl as a mobile phase, size exclusion chromatography (SEC) column removes the impurities by hydrophobic adsorption as well; molar masses obtained from both techniques thus agree when hydrophobic filters are used in bulk light scattering experiments. Diverse hydrodynamic flow retardation effects including slalom chromatography behavior are shown to substantially bias molar mass distributions obtained for ultra-high molar mass (UHM) HA, unless the flow rate in SEC analysis is reduced below 0.1 mL/min. Too high injected concentration (c inj ) is shown to introduce the onset of HA on column degradation. Correct polydispersity indices and molar mass distributions of UHM HA are obtained from SEC at a flow rate of 0.09 mL/min and optimized c inj .

Distribution analysis of cellulose acetate phthalate by ion-exclusion-moderated size exclusion chromatography

Abstract Ion-exclusion is the electrostatic repulsive interaction between a charged polymer and charges of the same sign on the surface of a column packing. Controlled ion-exclusion allows compensation of hydrophobic adsorption in size exclusion chromatography of negatively charged cellulose acetate phthalate (CAP) polymers in acetone/water/LiCl (80/20) as a mobile phase. Properly selected low-ionic-strength conditions provide correct separation in size-exclusion mode also in binary solvent mixtures. Possible interfering effects related to light scattering at low-salt conditions are shown to be negligible if on-line concentration/light scattering detection is used. The absence of these interferences is easily checked by a comparison of experiments at two different low-salt concentrations. Molecular weight averages and distributions identical within the experimental error are obtained when both salt concentrations are properly selected.

Flow irregularities due to polymer elution in size exclusion chromatography and its effect on viscometric detection

Abstract A new experimental setup for detecting flow-rate fluctuations caused by the passage of polymer through a separation system, including a differential refractometer and a singlecapillary on-line viscometric detector in size exclusion chromatography (SEC) is demonstrated. Numerical methods for correction of elution volume and the baseline of the viscometric output, including the “Lesec effect”, are presented on SEC data of (2-hydroxy-ethyl) cellulose in 50% methanol/50% water 10 mM NaCl (mobile phase). The effect of flow-rate fluctuations on experimental molecular weight averages is discussed.

Size distribution as a function of branching in the pregelation stage for poly(methyl methacrylate) containing randomly distributed tetrafunctional units

Size distribution in methyl methacrylate polymers containing small amounts of randomly distributed dimethacrylate units was studied as a function of branching up to the point of gelation. Size distribution was determined by size exclusion chromatography with refractive index and low angle light scattering detection as well as by dynamic light scattering. Proper handling of experimental data to get precise size averages is critically discussed. Multiangle static light scattering was used to provide additional verification of data obtained from size exclusion chromatography and dynamic light scattering. Expansion factors were determined for further elucidation of the effect of crosslinking in terms of coil volume change. The results agree quite well with the Flory-Stockmayer theory of gelation. However, the prediction of modern scaling theories that the ratio of the static and dynamic radii for randomly branched copolymers would attain a value of unity or slightly below as the critical branching, i.e. the gel point, is approached was not confirmed.