Calibration of gel-permeation columns in the high-molecular-mass range: fixed human thrombocytes for the estimation of interstitial volume and the haemocyanin of the Vineyard snail Helix pomatia as a molecular mass calibration substance

Barnikol, W. K. R.; Pötzschke, Harald

doi:10.1016/0021-9673(94)00690-3

Cited by 12 publications

(4 citation statements)

References 6 publications

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“…The void volume of the column, where the calibration curve should become vertical, with no further reduction of elution time with increasing molar mass, does not appear to have been reached. The value of the void volume for these columns is not clear; the problem of finding an appropriate polymer to measure void volumes has been described for aqueous SEC systems, where a protein extracted from snails has been used . We have shown elsewhere 1,27 that the polystyrene calibration predicts the elution times of standard molecules (polycyclic aromatics, azaarenes, oxygenated aromatics) up to 1000 u within about 1 min and defines the small molecule elution limit, the permeation limit, at not later than 23 min; no small molecule standards eluted after 22.7 min.…”

Section: Resultsmentioning

confidence: 99%

Size Exclusion Chromatography of Soots and Coal-Derived Materials with 1-Methyl-2-pyrrolidinone as Eluent: Observations on High Molecular Mass Material

et al. 2000

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A number of samples which eluted at unexpectedly short retention times during size exclusion chromatography have been characterized. Soot and tar samples likely to show similar behavior have been examined. Distinct peaks from about 6 min were observed, compared to 9−10 min for more usual samples. Molecular masses of the early eluting material appear to be large, although extrapolation of existing calibrations does not seem appropriate. A naphthalene mesophase pitch also gave peaks at short elution times. It appears reasonable to interpret chromatograms of this nonpolar material as a direct indication of the presence of large molecular mass material, and to infer that excluded peaks of SEC chromatograms do not necessarily result from the presence of clusters of polar molecules. GC-MS and probe-MS examination of the samples showed only very limited proportions of the samples to have small molecular masses. MALDI-MS spectra of the samples indicated the presence of signal up to 20 000 u. Taken together, data from SEC and the three MS techniques indicated the presence of very large molecular mass materials in this set of samples. Tar deposits recovered from entrained, combusting coal particles have also been examined, providing direct evidence for the presence of large molecular mass material in combustion environments. The observation contrasts with mathematical models of coal burners, where rates of combustion of volatiles are assumed similar to rates of combustion of methane. The nature of the early-eluting material is not known but repeated microfiltration and TEM indicate that it may correspond to molecular diameters in the region of 20 nm. The soots and other samples appear to be in true solution rather than in colloidal suspension.

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Section: Resultsmentioning

confidence: 99%

Size Exclusion Chromatography of Soots and Coal-Derived Materials with 1-Methyl-2-pyrrolidinone as Eluent: Observations on High Molecular Mass Material

et al. 2000

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“…The value of the “void” volume of the column, where the calibration curve would be expected to rise vertically with no further reduction of elution time with increasing molar mass, is not well-defined. The problem of finding an appropriate polymer to measure void volumes of SEC columns has been described for aqueous SEC systems, where a protein extracted from snails has been used . The effect of shear degradation on large molecules that elute near the void volume of the column caused by high flow rates of eluent has been shown .…”

Section: Resultsmentioning

confidence: 99%

Effect of LiBr Addition to 1-Methyl-2-pyrrolidinone in the Size-Exclusion Chromatography of Coal-Derived Materials

et al. 1998

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Two propositions relating to the interpretation of size-exclusion chromatograms (SEC) of coal-derived materials in 1-methyl-2-pyrrolidinone (NMP) have been examined. These were (i) that signal peaks showing up at exclusion (short retention time) limits of SEC columns are due to sample polarity alone and (ii) that shifts in SEC chromatograms to longer retention times, observed upon addition of LiBr to the eluent (NMP), are due to dissipation of ionic binding forces, causing disaggregation of polar clusters that would otherwise have appeared at retention times appropriate to larger molecular masses. In our experiments, effects due to polarity and molecular mass have been isolated by using two nonpolar samples (a naphthalene mesophase pitch and a mixture of fullerenes). In the presence of LiBr, precipitation of solute out of solution and shifts of chromatograms to longer retention times, unrelated to sample polarity, have been observed. A partial breakdown of the size exclusion mechanism was identified by the observed extension of chromatograms beyond the permeation limit of the column, similar to those observed when using eluents of insufficient solvent strength (e.g., THF, chloroform). Dosing LiBr into NMP sharply reduces the solvent power of NMP for coal-derived solutes. In the absence of LiBr, SEC chromatograms of the fullerene mixture, the naphthalene mesophase pitch, and its fractions separated by planar chromatography clearly showed significant signal under the “excluded” peak, entirely due to nonpolar material. The damage caused to the SEC column arising from precipitation of sample, in the presence of LiBr, was not permanent as had originally been feared. The balance of the evidence suggests that polarity of some molecules may cause shifts in their elution times to shorter values (larger apparent molecular masses) and that these may overlap with signal from large molecular mass material.

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“…The breakdown of the linear relationship for the highest mass standards may be clearly observed in Figure . The effective upper mass limit of the column for polystyrene standards has not been defined in this work; in aqueous systems, the problem of defining the void volume of SEC columns has been addressed by using a protein extracted from snails . The material eluting before the change of slope in Figure at 10 min is assumed to be excluded from the porosity of the column packing.…”

Section: Methodsmentioning

confidence: 99%

A Comparative Study of Bitumen Molecular-Weight Distributions

et al. 1999

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A sample of Athabasca bitumen was fractionated by preparative size-exclusion chromatography (SEC), and the molecular-weight distribution of the five fractions and the original sample were determined by SEC in NMP solvent using a calibration based on polystyrene standards, by 252 Cf plasma desorption mass spectrometry (PDMS), and by laser desorption and matrix-assisted laserdesorption mass spectrometry (LDMS and MALDI) and the average molecular weights were measured by vapor pressure osmometry (VPO). A comparison of the results shows that VPO, MALDI, and SEC gave similar results while PDMS showed the same trend. The molecular-weight distribution of this 525+ °C fraction is continuous up to and perhaps beyond a molecular weight of 15 000.

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Calibration of gel-permeation columns in the high-molecular-mass range: fixed human thrombocytes for the estimation of interstitial volume and the haemocyanin of the Vineyard snail Helix pomatia as a molecular mass calibration substance

Cited by 12 publications

References 6 publications

Size Exclusion Chromatography of Soots and Coal-Derived Materials with 1-Methyl-2-pyrrolidinone as Eluent: Observations on High Molecular Mass Material

Size Exclusion Chromatography of Soots and Coal-Derived Materials with 1-Methyl-2-pyrrolidinone as Eluent: Observations on High Molecular Mass Material

Effect of LiBr Addition to 1-Methyl-2-pyrrolidinone in the Size-Exclusion Chromatography of Coal-Derived Materials

A Comparative Study of Bitumen Molecular-Weight Distributions

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