Handbook of Size Exclusion Chromatography and Related ...

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Table 7 Analytical Approach for Starch-Glucan Characterization and Parameters that may be Obtained Approach Experimental Parameter Information Preparative SEC; purification þ pooling Total hydrolysis þ anthron coupling ! E 540 (total carbohydrates); iodine staining: E 525, E 640 Fragmentation Step by step fragmentation; ! pure chemical; ! enzymatically catalysed; fragment analysis Semipreparative SEC; in-line elution profile; off-line complexing Analytical SEC: mass detection þ molecular weight calibration Analytical SEC: mass/ LALLSdetection Analytical SEC: mass/ viscosity detection © 2004 by Marcel Dekker, Inc. In-line DRI-detection offline iodinecomplexing þ E 640, E 525 detection In-line DRI detection ! mass_ev Elution profiles: DRI ! mass_ev LALLS ! LS_5_EV Elution profiles: DRI ! mass_ev visc ! eta_spec E 540 ! mass (V ret) E 640/E 525 (V ret) Type of fragments; mol of fragments; mass of fragments; Mol (V ret) mass (V ret) E 640/E 525 (V ret) Mass (Vret) concentration dn/dc Mass (V ret) R Q (V ret) M (V ret) Mass (V ret) [h] (V ret) V e (V ret) Mass fractions distribution; lcb/scb ratio Constituting glucan distribution; mean glucan conformation; branching characteristics: ! chain lengths of branches ! br% (percentage of br.) ! # of branching positions Mass fractions distribution; molar fractions distribution; lcb/scb ratio Mass fractions distribution; molar fractions distribution; recovery info on preferential dissolution Apparent absolute molecular weight calibration (log(M) vs. Vret) ! Mn, dpn/Mw, dpw ! m_MWD_d, m_dpD_d ! n_MWD_d, n_dpD_d Excluded volume Vn distribution ! m_VeD_d, n_VeD_d
Table 7 (Continued) Approach Experimental Parameter Information Analytical SEC: mass/molar detection Analytical SEC: mass/LS/ visc detection Bulk viscosity h below overlapping concentration c* Bulk viscosity h at varying mechanical stress (shear deformation D) Bulk viscosity h above overlapping conc c* at varying thermal stress (T) Bulk viscosity h above overlapping conc c* at varying thermal stress (T) and const. mechanical stress Photon correlation spectroscopy (PCS) © 2004 by Marcel Dekker, Inc. Elution profles: DRI ! mass_ev UV/VIS/ fluorescence ! mol_ev Elution profiles: DRI ! mass_ev LS ! LS_u_EV visc ! eta_spec Viscosity of concentration series Complex viscosity at varying shear deformation; for dissolution periods Viscosity at increasing temperature Brabender viscosity: constant shear deformation period of incr. T period of holding elev. T period of decr. T Translational diffusion coefficient DT Mass (V ret) mol (V ret) M (V ret) Mass (Vret) RQ (Vret) M (Vret) [h] (Vret) Ve (Vret) h(c!0) ; [h] av c* av ¼ 1/[h] av h* (o) h* (D) (t) T dis DT dis Viscosity as function of temperature, deformation and application time: h(T, D, t) D T ! R H ! l coh Distribution of mass fractions ! m_D TD Distribution of LS-intensity fractions ! intensity_D TD Absolute molecular weight distribution ! Mn, dpn/Mw, dpw ! m_MWD_d, m_dpD_d Universal SEC calibration ! VeD_d Mean value of excluded volume [h] av; overlapping concentration c* av Visco-elastic properties of starch glucan solutions: ! Newtonian/ non-Newtonian Conformational stability; gelatinization temperature; disintegration temperature Stability/resistance towards applied energy (T, D, t): ! disintegration characteristics ! re-organization capability Population analysis: translational diffusion coef. DT; sphere equivalent radii of diffusing molecular objects RH; coherence length of molecular and supermolecular segments
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MARCEL DEKKER Handbook of Size Excl
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CHROMATOGRAPHIC SCIENCE SERIES A Se
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60. Modern Chromatographic Analysis
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Preface to the Second Edition Gel p
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Preface to the First Edition Molecu
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Contents Preface to the Second Edit
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17. Size Exclusion Chromatography o
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James F. Curry Analytical Departmen
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Iwao Teraoka, Ph.D. Othmer Departme
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The graphical data display typicall
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polymer molecule can elute. The tot
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individual states) allowed to them.
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unsaturation while the IR detector
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Sampleconcentrationshouldbeminimize
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averagesdefinedintermsofthemolecula
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information regarding appropriate M
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Analytical. Two models, the KMX-6 a
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Figure 6 Overlay of time-sliced pea
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accurately for very large and very
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4 GENERAL REFERENCES The interested
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46. TA Chamberlin, HE Tuinstra. US
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materials are commercially availabl
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Table 1 Commercial Column Packing M
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necessarily comparable. For this re
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adjusted such that the pore size di
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narrowmolecular weight range, indiv
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Figure 8 Effect of particle size on
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Figure 9 SEC calibration using poly
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Table 4 Typical Eluent Systems for
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Figure 12 Analysis of poly-2-vinyl
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24. E Meehan, S O’Donohue. The ro
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into accepted laboratory techniques
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Table 1 Trace Metal Impurities in C
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mass and biological activity, for m
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Experimental efficiency vs. velocit
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Table 3 Properties of SEC Silica Ge
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operate at aflow rate that can easi
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Figure 3 Efficiency of amide-bonded
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mechanically stable packing materia
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Figure 5 Pore size distributions of
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Figure 6 Separation of polystyrenes
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60-120 A ˚ are large enough to be
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Table 6 Selected Silica-Based Colum
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Table 7 Separation Ranges for Polym
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the decline in permeability is perm
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sample components can be varied by
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Figure 10 Diol bonding reactions. I
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ecause of the higher relative molec
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valuesfork1,k2,anda,theunknownmolec
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water-soluble and detergent-soluble
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the importance of secondary retenti
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discussed in Table 8(33). Based on
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individual dispersion processes ins
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The detector time constant can dist
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5.3 Mobile Phase A mobile phase is
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less steep and still linear at high
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35. P Bristow, J Knox. Chromatograp
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112. K Gooding, K Lu, G Vanecek, F
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for branched polymers or copolymers
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Figure 1 “Bridge design” flow-r
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Figure 3 A light-scattering photome
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Figure 5 SEC universal calibration
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where hn is the hydrodynamic volume
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It should be noted that dn=dc also
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sensitivity for many samples compar
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that is, log[h] vs. logM,generated
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Figure 8 Effect of band broadening
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Table 1 Measurement of Molecular We
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Figure 10 Differential refractomete
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Figure 11 (A) Mark-Houwink plot of
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Table 3 Measurement of Molecular We
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size information is derived from un
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Table 5 Molecular Weight Distributi
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By measuring the scattered intensit
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8 APPENDIX: INSTRUMENT COMPANIES 8.
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53. G Marot, J Lesec. J Liq Chromat
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125. D Slootmaekers, JAPP Van Dijk,
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200. JG Bindels, GJJ Bessems, BM De
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possibly with hydrodynamic volume (
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An insurmountable limitation of SEC
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to sDR(V) when either nPS ¼nPB ¼c
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1,4-trans;and8%1,2-vynil.TheglobalC
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The goal is to find the MWD and CCD
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As expected, pS is closer to the no
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Figure 3 Example 3: MWD and BD of a
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9. BH Zimm, WH Stockmayer. The dime
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45. RO Bielsa, GR Meira. Linear cop
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solubility properties. They are use
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determination of the absolute molec
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Figure 4 Result of the PBT analysis
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Figure 8 Result of natural spider s
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7 Size Exclusion Chromatography of
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As pointed out earlier, rubber must
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Table 2 Various Solvents and Operat
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Figure 2 Typical GPC calibrations w
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© 2004 by Marcel Dekker, Inc. Figu
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Natural and Synthetic Rubbe177 vari
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Another example, shown in Fig. 5 (3
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APPENDIX: SEC CONDITIONS FOR RUBBER
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Appendix (Continued) Polymer Column
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5. J West, E Rodriguez. Rubber Chem
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associate. Girdler (67) and Speight
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Table 1 Fractions Obtained Using Co
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Figure 1 SEC analyses of an aged as
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Apparently more polar compounds are
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40% was fractionated into four frac
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Figure 7 SEC analyses of an unaged
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Figure 9 Comparison of percentage L
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Figure 11 Comparison of SEC chromat
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Figure 12 Comparisons of SEC chroma
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percentage LMS at the other locatio
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the total area into up to 12 sectio
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attempted to develop an SEC method
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parameterisbasedontheinternalpressu
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5 MODIFIED ASPHALTS The addition of
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Oxidation of the rubber and asphalt
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Figure 20 SEC chromatogram for an a
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Figure 21 Comparisons of apparent m
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© 2004 by Marcel Dekker, Inc. S/
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PS/10 4 þ 10 3 þ 500 þ 100 THF/1
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Nevertheless, SEC of asphalts is es
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56. NW Garrick, RR Biskur. Trans Re
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Asphalts 235 122. M-S Lin, JM Chaff
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Table 1 Applications of Acrylamide
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(AM/AA), and acrylamide/dimethyldia
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polymer, a large size filter of 5,
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Figure 1 Size exclusion chromatogra
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Table3 MWandMWDofaBroad-MWDPAMStand
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Another commercially available MW d
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Table 5 Weight-Average Molecular We
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Figure 6 (a) Size exclusion chromat
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compared to anormal product. By com
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StudyingtheKineticsofaChemicalReact
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this information, the higher MW pea
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46. CJ Kim, A Hamielec, A Bendek. J
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A relatively new technique utilizin
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where ais the exponent of the Mark-
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Figure 2 TDS chromatograms for full
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Figure 5 Comparison of molecular we
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Table 3 Summary of TDS Molecular We
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thepartiallyhydrolyzedPVAwiththecol
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Table 5 Summary of Conformation and
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a¼0.627andlog(K) ¼23.249.Theseare
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The limits of detection of the PVA
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Figure 12 PVAc broad molecular weig
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4 SUMMARY Advances in SEC character
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2.1 Molecular Weight Grades of PVP
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exclusion chromatography with low-a
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weightandmolecularweightdistributio
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Figure 4 PEOcalibrationofUltrahydro
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Figure 6 Overlay of GPC chromatogra
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plus Ultrahydrogel 120 A ˚ ,Shodex
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Figure 10 SEC traces of PVP/AA copo
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Figure 11 Molecular weight distribu
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2 CHEMICAL, MACROMOLECULAR AND MORP
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Table 2 The Relative Composition of
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considered to be 3. After the DS ha
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een extensively studied by methods
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The silylation was performed in LiC
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observed using column materials hav
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Table 6 (Continued) Packing materia
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4.5 Other Cellulose Derivatives Bes
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Table 8 SEC Conditions for Characte
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dextrans swell too much in cadoxen
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stirring at room temperature for an
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Figure 2 MMDofunbleached(HP)andblea
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Figure 4 MMDofbirchkraftpulpdegrade
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adequate when evaluating the influe
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differential refractive index (DRI)
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30. HA Krässig. Effects of structu
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67. E Sjöholm, K Gustafsson, A Col
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104. B Fleury, J Dubois, C Léonard
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136. D Miller, D Senior, R Sutcliff
Page 367 and 368: 166. SS Cutié, CG Smith. Determina
Page 369 and 370: 200. R Berggren, F Berthold, E Sjö
Page 371 and 372: 13 Molar Mass and Size Distribution
Page 373 and 374: measurements, and vapor pressure os
Page 375 and 376: The calibration of SEC columns by c
Page 377 and 378: The division according to eluent ty
Page 379 and 380: Apolymer produced by UV irradiation
Page 381 and 382: Because DMAC/LiCl is also agood sol
Page 383 and 384: compoundsandwasexplainedbyadsorptio
Page 385 and 386: isolated lignins because they are e
Page 387 and 388: The Separon HEMA and Separon HEMA B
Page 389 and 390: Figure 4 GPC elution curves of the
Page 391 and 392: liquor is highest throughout the co
Page 393 and 394: topological anisotropy in the polym
Page 395 and 396: 29. M Ristolainen, R Alen, J Knuuti
Page 397 and 398: eds. Proc Int Symp 1998. Canton, Pe
Page 399 and 400: 92. L Jurasek. Towards a three dime
Page 401 and 402: Table 1 Selected Industries Connect
Page 403 and 404: into C3-metabolites, which then may
Page 405 and 406: Table 4 Key Enzymes in the Biosynth
Page 407 and 408: Table 5 Cloned Mutants of Starch En
Page 409 and 410: Figure 3 Modeled x-ray diffraction
Page 411 and 412: In the native environment (plant ce
Page 413 and 414: Figure 6 (a) Large starch granules
Page 415 and 416: number of branching points); crossl
Page 417: Both absolute approaches are extrao
Page 421 and 422: structures, excluded volumes, and t
Page 423 and 424: Figure 10 (a) SEC elution profile o
Page 425 and 426: Figure 12 Wheat glucans: DRI elutio
Page 427 and 428: Figure 13 Wheat glucans. Normalized
Page 429 and 430: Figure 16 Wheat glucan. Normalized
Page 431 and 432: Figure 18 Wheat glucan.Elution prof
Page 433 and 434: Figure 20 Wheat glucans. Absolute m
Page 435 and 436: Figure 22 Wheat glucan. (a) Intrins
Page 437 and 438: Figure 24 Wheat PA-glucan. Elution
Page 439 and 440: an Ubbelohde-viscometer for concent
Page 441 and 442: Rheological investigations of stabi
Page 443 and 444: shifts to applied laser light, whic
Page 445 and 446: Table 8 Characteristic Parameters f
Page 447 and 448: Table 8 (Continued) and H2O. A part
Page 449 and 450: 10. SG Ball, MHBJ van de Wal, RGF V
Page 451 and 452: 52. W Praznik, R Beck. J Chromatogr
Page 453 and 454: 15 Size Exclusion Chromatography of
Page 455 and 456: 3 PROTEIN PARTITIONING IN SEC 3.1 G
Page 457 and 458: Figure 1 Plot of Kav vs. log M for
Page 459 and 460: yconsideringtheproteinsolutestobesp
Page 461 and 462: globular proteins and selected flex
Page 463 and 464: silanol groups (52-54); even capped
Page 465 and 466: most proteins to be maximally stabl
Page 467 and 468: Figure 4 Chromatograms of BSA and P
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concentrated) steps of protein puri
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Table 3 Characteristics of Dextran-
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preparative SEC. The capabilities o
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37. GR Noll, N Nagle, JO Baker, DJ
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Figure 2 Separation of total E. col
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Figure 3 Separation of HaeIII-cleav
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The recovery of DNA fragments has b
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Endotoxin should also be removed fr
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Table 3 Best Columns for the Separa
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Figure 10 Dependence of HETP on flo
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Appendix (Continued ) Polymer Colum
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REFERENCES 1. CT Wehr, SR Abbott. J
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MALDI-MS has been applied to determ
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Figure 1 Calibration curvesof SEC c
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Figure 3 Chromatograms of epoxy res
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wherenandn0 aretheRIsofthesample an
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Figure 5 SEC chromatogram of n-alka
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Figure 7 Refractive indexes of comm
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Another detector recently applied i
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Figure 11 Absolute MW calibration p
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Figure 12 Chromatogram of Acrawax C
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18 Two-Dimensional Liquid Chromatog
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largerispressuredropandresultingexp
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elations between molar masses and s
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Figure 2 (Continued) chapter, one o
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where Nis the column efficiency exp
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composition/architecture/molar mass
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solvent. Here again, solvent-solven
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or repulsive interactions between m
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macromolecules may strongly differ
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temperature. The efficacy of an ads
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however, aliphatic bonded groups ma
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Figure 10 Schematic representation
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packing materials for SEC of synthe
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Quantitative relations between sila
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Both strength and thermodynamic qua
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where Cand Dare constants for apart
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ThisisimportantwhenLCCCisappliedtoc
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Page 549 and 550:
Page 551 and 552:
However, strong adsorption of macro
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elution step may be sufficient for
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Figure 16 Block scheme of a full ad
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processing of 2D-HPLC data. Knowled
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Figure 20 Set of full retention-elu
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distributions of complex polymers a
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constituents were not discriminated
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(Sec. 7) and therefore they can be
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2. Identify sample solvents. First
Page 569 and 570:
1 Segmental interaction energy para
Page 571 and 572:
58. BG Belenkii. Pure Appl Chem 51:
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19 Methods and Columns for High-Spe
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Table 1 Summary of Methods for Incr
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not savealot of time and solvent, d
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Figure 3 Conventional SEC chromatog
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chromatogram at 4mL/min, which is f
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Sincethereductionoftheinternaldiame
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Table 3 Summary of Column Performan
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Figure 11 Dependence of column perf
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. Two-dimensional chromatography ru
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The standard deviations for the mol
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Figure 15 Comparison of time requir
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Figure 17 Accuracy and precision of
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polymer analysis. Samples are “ru
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just ignoring the fractionation of
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20 Automatic Continuous Mixing Tech
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out at low enough concentration tha
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educed viscosity, h r, to be comput
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Page 609 and 610:
Figure 2 Raw ACOMP signals from mul
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Figure 4 Mw vs. monomer conversion,
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decreases smoothly and monotonicall
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Figure 7 Effects of fluctuating rea
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Finally, the use of a light-scatter
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Figure 8 Linear and cubic increases
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where r(t) is now given by 2 6 r(t)
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Figure 11 Light scattering from sol
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3.1 ASimple System: Equilibrium Cha
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Figure 13 (a) ACM applied to the ch
Page 629 and 630:
Also of note in Fig. 13b is how hig
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15. JM Starita, CL Rohn. Rheologica
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52. M Hulden. Hydrophobically modif
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21 Light Scattering and the Solutio
Page 637 and 638:
therefore, is to remove any lingeri
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is often found in the literature wh
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Figure 2 Configuration of an SEC se
Page 643 and 644:
5 THE IMPORTANCE OF SOFTWARE Like a
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standard deviations of the measured
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These include the differential weig
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Figure 8 Data of Fig. 7corrected fo
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Figure 11 Data of Fig. 10 with spik
Page 653 and 654:
Figure 13 Fitstodatacollectedatasli
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Table 1 Errors (in %) Characteristi
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homogeneous copolymer, that is, tak
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process may be used to identify the
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fractionated sample. They represent
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Figure 16 An example of poor SEC ch
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on) methods, whereby the average di
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23. WH Stockmayer, LD Moore Jr, M F
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condition, as is explained in this
Page 671 and 672:
Figure 2 Partition coefficients KL
Page 673 and 674:
4 COLUMNS FOR HOPC Detailsaregiveni
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and concomitant clogging of columns
Page 677 and 678:
Figure 5 Concentration dependence o
Page 679 and 680:
Table 1 Solution Volume, Polymer Ma
Page 681 and 682:
in Fig. 4a. The middle fractions (8
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Figure 10 SEC chromatograms for som
Page 685 and 686:
appropriate for the early fractions
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23 Size Exclusion/ Hydrodynamic Chr
Page 689 and 690:
Figure 2 Calibration curve for the
Page 691 and 692:
Figure 5 Elutionbehaviorofpolystyre
Page 693 and 694:
Table 1 Comparison Between SEC and
Page 695 and 696:
egular SEC column in which the pore
Page 697:
Such an ideal separation can be obt
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