Non-porous Glass Beads Research Articles

Flow-rate effect on elution volume in size-exclusion chromatography

For years it has been noticed that elution volume can change with flow-rate in size-exclusion chromatography (SEC). However, the observations are inconsistent and the mechanism is still unclear. The present study conclusively demonstrates the flow-rate effect on elution volume. Two porous silicas with quite different pore structures and two non-porous glass beads with very different sizes are carefully examined at various flow-rates. Benzene and polystyrenes are employed as the probe solute with dichloromethane as the eluent. The elution is performed on an isocratic high-performance liquid chromatography system equipped with a computer. The flow-rate is measured by a pipet in which a thin water layer is placed to minimize evaporation of the effluent. The flow-rate is approximately averaged on the retention time period to achieve the best accuracy. The retention time is calculated from the first moment using a BASIC program. The experimental results clearly show that elution volume increases linearly with an increase in flow-rate for all solutes in all packings. The ratio of slope to intercept is defined as the ascending rate for mutual comparison. Consistent with previous reports, the ascending rate increases with an increase in the probe solute size for all columns. Furthermore, the ascending rates for two non-porous packings are much greater than those for porous packings. This reveals that hydrodynamic volume is probably the primary factor responsible for the flow-rate effect on elution volume and the stagnant volume in pores does not contribute to the flow-rate effect. This inference is consistent with the previous hypothesis of “hydrodynamic induced diffusion”. However, it should be emphasized that flow-rate would not affect the SEC distribution coefficient significantly and its effect can be neglected in practice.

9] Covalent immobilization of proteins by techniques which permit subsequent release

Publisher Summary Characterization of immobilized species has been restricted to kinetic studies or to examination of fluorescent spectra because of the limitations imposed by the immobilization matrix. Among the approaches that have been used are derivatizations that permit cleavage of the immobilized species from the matrix with aqueous hydroxylamine and those that permit cleavage through the reduction of disulfide bridges. This chapter discusses thionyl chloride-activated succinamidopropyl-glass as an immobilization matrix. Succinamidopropyl-glass can be prepared from porous or nonporous glass beads by either an aqueous or a nonaqueous derivatization procedure. The aqueous procedure results in a surface that is more hydrated and contains more silane polymers than that produced by the nonaqueous method. Although both procedures yield surfaces that will covalently immobilize proteins (or other suitable nucleophiles) following activation with thionyl chloride, and that will subsequently release immobilized proteins upon treatment with hydroxylamine, the fact that subtle differences in the surfaces are obtained should be borne in mind in selecting the best method for a specific application.

Characteristics of Pore-Size Distributions of Sandstone Reservoir Rocks and Their Relationships to Permeability: ABSTRACT

Cumulative pore-size distribution curves of sandstones determined by mercury porosimetry always consist of a few segments of normal Gaussian distribution when they are drawn on a log-probability diagram. Numerous mechanically packed assemblages were prepared to elucidate characteristics of segmentations from nonporous glass beads and porous shale fragments which represent constituent mineral grains and rock fragments of sandstones, respectively. These assemblages are classified into three groups. (1) Assemblages composed of sand-size glass beads. In addition to equal-size assemblages (1, 1.5, 3, 5 phi) and unequal-size assemblages (sorting is 0.42, 0.64, 0.89), assemblages of angular quartz grains are also examined to know the effect of angularity of grains. (2) Assemblages of sand-size and matrix size beads. Matrix-size beads are arranged in conformity with normal distribution in size. (3) Assemblages of sand-size beads and shale fragments. Proportions of shale fragments to beads are varied to define the effect of intragranular porosities of shale to total porosities. According to these sphere-packed models, segments of normal distribution may be explicitly explained by (1) a framework distribution of intergranular pores built up by grain packing followed by a subordinate distribution of toroidal voids around grain contacts, (2) smaller size intergranular pores by matrix-size grains, and (3)more » intragranular pores contained in rock fragments. The same conclusions are obtained in Tertiary sandstones from the back arc of Japan. Permeability of sandstones may significantly suffer from a framework distribution of intergranular pores built up by grain packing, and specific surface area may also prescribe permeability in addition to porosity and pore size.« less

Analytical high-performance affinity chromatography: evaluation by studies of neurophysin self-association and neurophysin-peptide hormone interaction using glass matrices.

Bovine neurophysin II (BNP II) was covalently immobilized on both nonporous and porous (200-nm pore diameter) glass beads and incorporated in a high-performance liquid chromatograph to evaluate analytical high-performance affinity chromatography as a microscale method for characterizing biomolecular interactions. By extension of the theoretical treatment of analytical affinity chromatography, both the self-association of neurophysin and its binding of the peptide hormone vasopressin were characterized by using a single chromatographic column containing immobilized neurophysin predominantly in the monomer form. Both [3H] [Arg8]vasopressin (AVP) and 125I-BNP II were rapidly eluted (less than 25 min). The relatively symmetrical elution peaks obtained allowed calculation of both equilibrium dissociation constants and kinetic dissociation rate constants. The dissociation constant measured chromatographically for the AVP-immobilized neurophysin complex, KM/L = 11 microM with porous glass beads and 75 microM with nonporous glass (NPG) beads, was in reasonable agreement with those previously obtained by curve fitting of Scatchard plots (16-20 microM) and from binding to [BNP II]Sepharose (50 microM). The values obtained are larger than that for dissociation of AVP from BNP II dimer, by a factor consistent with the intended nature of immobilized BNP II as monomers. Chromatography of BNP II on the [BNP II]NPG gave a dimer dissociation constant of 166 microM, a value in excellent agreement with that derived from equilibrium sedimentation studies (172 microM). In contrast to the agreement of chromatographic equilibrium binding constants with those measured in solution, the dissociation rate, k-3, determined from the variance of the affinity chromatographic elution profile with nonporous beads, was several orders of magnitude smaller than the solution counterpart.(ABSTRACT TRUNCATED AT 250 WORDS)

Hydrodynamic studies in large-diameter columns

Axial dispersion resulting from the flow pattern alone has been investigated in two columns with internal diameters of 8 and 60 mm. To eliminate interactions between the solute and the support, a bed of non-porous glass beads was used, with sulphuric acid as a tracer. The solute concentration was measured at various points inside the column by conductivity probes. On the smaller column, the mean values of the concentration over the column cross-section were observed, in addition to local values at the centre of the packing, at two locations along the central line. On the larger column, local values were determined at two locations along the central line and at different points on a diameter; external cells were also located at the column inlet and outlet. The axial dispersion was evaluated with the dispersed plug-flow model, by comparing the conductivity signals at the two positions in the column. Various results are presented and compared with existing models: the dispersion in the two columns is compared; on the larger column, the local dispersion in the packing is explored and the contributions to dispersion caused by the packing and the column heads are evaluated.

Determination of macropore diffusion in molecular sieve particles by pulse gas chromatography

The experimental conditions necessary for reliable determination of the intraparticle diffusivity of gases in porous solids by pulse gas chromatography (PGC) are investigated. Diffusivity measurements carried out with non-adsorbable tracer gases in packings of molecular sieve particles of different sizes indicate that the experimentally observed contributions of external mass transfer and intraparticle diffusion resistance may not depend on the particle size as theory predicts. With decreasing particle diameter the external mass transfer resistance becomes increasingly dominant as compared to intraparticle diffusion resistances. This finding follows from comparative experiments with packings of non-porous glass beads and porous molecular sieve particles of equal size. The sensitivity of the mass transfer rate parameter determination is markedly improved by using an experimental arrangement with a column to particle diameter ratio which corresponds to a “single pellet string reactor” (S.P.S.R.). The realibility of the determination of the intraparticle diffusivity with an S.P.S.R. has been checked by independent diffusivity measurements carried out on pressed single pellets of the molecular sieve. Steady-state counter-diffusion and dynamic pulse response measurements yielded an effective diffusivity comaparable to that determined with the PGC method using an S.P.S.R. arrangement.

Immobilization of glucoamylase from Aspergillus niger on poly(ethylenimine)-coated non-porous glass beads

Glucoamylase (1,4-α- d-glucan glucohydrolase, EC 3.2.1.3) from A. niger was immobilized on cationic nonporous glass beads (13–44 μm) by electrostatic adsorption followed by rosslinking with glutaraldehyde. Over 80% of the enzyme's total soluble activity was expressed upon immobilization. d-Glucose production from maltodextrins was virtually complete, suggesting that the lack of pores can eliminate the problem of product reversion. Immobilized glucoamylase showed decreased stability upon heating, compared with the soluble enzyme .

Fixed-time kinetic enthalpimetry: improved sensitivity for enthalpimetric enzyme activity determinations in homogeneous and heterogeneous systems

A fixed-time (integral) method is described for the enthalpimetric determination of enzyme activity. The method involves the determination of residual unreacted substrate after a fixed incubation time with the sample. Results are presented for the determination of cholinesterase in aqueous solution and in 0.1cm 3 samples of reference sera using 30-min incubation periods. Results are correlated with a spectrophotometric procedure. A precision of 1.8% relative standard deviation is reported for serum assays. Preliminary data are also presented for the enthalpimetric determination of cholinesterase activity after immobilization onto non-porous glass beads by carbodiimide and glutaraldehyde coupling procedures.

Effects of immobilization on the kinetics of enzyme-catalyzed reactions. II. Urease in a packed-column differential reactor system.

Urease from Jack bean was immobilized on nonporous glass beads by covalent bonding and its kinetics were studied in a packed-column differential reactor. To facilitate comparison, the urease was immobilized by both diazo and glutaraldehyde coupling. The kinetic properties of immobilized urease were similar to those of the soluble enzyme and different immobilization methods did not appreciably alter the kinetic properties. The affects of three different amino acid activators appear to follow predictions obtained from a relatively simple competitive model, except at very low substrate levels.

Axial dispersion in beds of small particles

Axial dispersion coefficients (E A) in beds of small (0.1–1 mm) particles are often needed to interpret reaction rate data. A chromatographic technique was used to evaluate such coefficients for beds of porous catalyst particles and beds of non-porous glass beads over a wide range of velocities (Re p = 0. 00237 to 11.9). Two gaseous systems (helium in hydrogen and oxygen in nitrogen) were used to provide different molecular diffusivities. The results can be correlated by adding two terms: one due to molecular diffusion, which is dominant at low velocities, and a second term due to convective mixing, which is dominant at high velocities. The unusually high values of E A for porous particles at low velocities could be explained by including a contribution due to intraparticle diffusion. At high velocities Peclet numbers approached constant values which were proportional to particle diameter. Channeling in the bed due, for example, to shrinkage of catalyst particles as a result of reduction in situ, or to movement of particles, can cause a large increase in axial dispersion.

Separation Mechanisms in Gel Permeation Chromatography

Abstract This paper presents flow rate studies, vacancy chromatography, and a static mixing experiment. Data obtained on an unpacked column (a straight tube) and on a column packed with nonporous glass beads are also reported. The results reveal that peak dispersion in GPC arises mainly from lateral diffusion in the stationary phase (permeation in and out of the porous substrate) and from lateral diffusion in the mobile phase. GPC peak separation is mainly dominated by the process of steric exclusion. Pore size distribution data obtained on Bio-Rad porous glass are shown to illustrate the preference of random coil theories over theories of the equivalent sphere in the interpretation of steric exclusion of flexible polymers. The data are discussed in terms of Herman's diffusion theory and Cassasa's exclusion theory.

Extraparticle diffusional effects in gel permeation chromatography. II. Experimental results

AbstractExperiments coupled with a systems analysis were conducted on chromatogram dispersion, or zone broadening, in gel permeation chromatography (GPC). Three components of a Waters Associates Model 200 chromatograph, each of which is a potential cause of dispersion, were considered; these are: the packed column, with extraparticle dispersion only; the empty tubing, between pump and columns, columns and detector, etc.; and the detection system, viz., the differential refractometer cell. Toluene solvent was used and the solutes whose dispersion was studied included orthodichlorobenzene (ODCB) and narrow‐molecular‐weight polystyrene standards having molecular weights of 900, 20,400, 51,000, 97,200, and 160,000. Nonporous glass beads, 50 μ in diameter, were used as column packing. Two diameters, 1 mm and 0.5 mm, of stainless steel tubing were studied. In addition to the usual rectangular pulse sample injection, a step input mode for solute introduction was also used. The empty tubing was found to contribute significantly to the degree of dispersion and to skewness of elution curves. Anomalous bimodal characteristics of the elution curves were also observed which could only be ascribed to the empty tubing. These phenomena depended markedly on parameters such as tube diameter and length, and solute concentration and molecular weight. Dispersion in the packed column, although important, was found to be symmetrical (Gaussian) and less sensitive to these parameters than in the empty tubing, especially with respect to molecular weight. Dispersion in the cell was believed to be insignificant relative to the packed column and empty tubing.

Evaluating dispersion in gel permeation chromatography. Axial dispersion of polymer molecules in packed beds of nonporous glass beads

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTEvaluating dispersion in gel permeation chromatography. Axial dispersion of polymer molecules in packed beds of nonporous glass beadsR. N. Kelley and F. W. BillmeyerCite this: Anal. Chem. 1969, 41, 7, 874–879Publication Date (Print):June 1, 1969Publication History Published online1 May 2002Published inissue 1 June 1969https://doi.org/10.1021/ac60276a006RIGHTS & PERMISSIONSArticle Views101Altmetric-Citations33LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (638 KB) Get e-Alerts Get e-Alerts