Gel permeation, chromatography,

Gel permeation ehromatography was run in 2,5-dichlorotrifluoromethylben-zene at 130°C. The results of samples from different sources are shown in Table 

Example Initiator (%) Solvef Temperature (°C) Yield (%) Inherent viscosity (dl/g CTsCl) 

Example Sample Zero strength time Inherent viscosity (dl/g C6F5CI) Mn My, P 

Conditions eluent 2,5-dichlorotrifluoromethyl benzene, temperamre 130°C (Run by Mike Stephens, CRL 

The nonionic gel stationary phase is commonly composed of cross-hnked polystyrene or macroporous silica particles, which do not swell significantly in the carrier solvents. A range of pore sizes is fundamental to the success of this size fractionation procedure, which depends on two processes. These are (1) separation by size exclusion alone, which is the more important feature, and (2) a dispersion process, controlled by molecular diffusion, which may lead to an artificial broadening of the MMD. 

We consider first the mechanism of the separation process in simple terms, the large molecules, which occupy the greatest effective volume in solution, are excluded from the smaller pore sizes in the gel and pass quickly through the larger channels between the gel particles. This results in their being eluted first from the column. 

The efficiency of the separation process is then a function of the dependence of the retention (or elution) volmne Vj on the molar mass M, and a rehable relationship between the two parameters must be estabUshed. The value of depmds on the interstitial void volume Vq di accessible part of the pore volume in the gel. 

To obtain the MMD, the mass of the polymer being eluted must be measured. This can be achieved continuously using refractive index (RI), UV, or IR detectors, which will give a mass distribution as a function of V. It is still necessary to estimate the molar mass of each fraction before the MMD curve can be constructed. If the universal calibration curve is valid for the system, then 

FIGURE 9.9 Elution curve schematically showing the range of elution volumes that are valid for a particular column. In this case, molecules with molecular size tP are totally excluded and eluted without discrimination, whereas those tend to become absorbed or are partitioned if a mixed solvent is used and Kd= 1. 

Gel filtration chromatography using Sephadex G100 as column packing and ultraviolet detectors, have been used in studies carried out on the elution of humic acid [8] and in characterisation studies on secondary sewage effluents [9] and in organic substances in river waters [10], 

McKay and Lathan [11] used gel permeation chromatography to determine polyaromatic hydrocarbons in non saline waters. 

Done and Reid [12] applied gel permeation chromatography to the identification of crude oils and products isolated from estuary and sea water, The technique, which appears more suited to the analysis of crude oils, is based on the separation of oil components in order of their molecular size, for practical purposes their molecular weight. 

Collins and Webb [13] used gel chromatography to detect and determine carbohydrates in pulp mill effluents. The phenol-sulphuric acid method was used to monitor column effluents. 

Sagfors and Starck [14] used gel permeation chromatography to study calcium lignosulphate of high molecular weight in acid and alkaline kraft pulp bleaching effluents. 

Gel permeation chromatography (GPC) is essentially a process for the separation of polymer molecules according to their size. The separation occurs as the solute molecules in a flowing liquid move through a stationary bed of porous particles. The method has been used extensively in biochemistry to separate biological polymer molecules from small molecule contaminants (with the use of Sephadex column). Application of the method to synthetic polymer chemistry in the 1970s has revolutionized the procedures for polymer characterization and molecular weight determination. 

The principle that underlies the method can be explained as follows Assume that a dilute solution containing a broad molecular weight distribution of polymer chains and oligomers is allowed to flow through a column packed with finely divided solid particles, which have pores (tunnels) of diameter, say, 1000 X. As the dissolved solute passes each particle, molecules with dimensions smaller than 1000 X will enter the pores and will permeate the pore space under the influence of the usual thermal 

the GPC method is essentially a process for the fractionation of polymers according to their size, and hence according to their molecular weight. However, the molecular weight cannot be determined directly. Molecular weight can be calculated from the GPC data only after calibration of the GPC system in terms of retention time or elution volume with polymer standards of known molecular weights (see p. 298). 

A schematic layout of a typical GPC unit is shown in Fig. 4.24. Since the elution rates by gravity flow through a vertical column, as in conventional chromatography, will be slow and nonreproducible, a mechanical pump is usually employed to force the sample and the elution solvent through the column at pressures of up to 1000 to 4000 psi and at a rate of 2 to 3 ml/min. The sample is injected into the column entry from a graduated hypodermic syringe (typically, 0.5 to 3 ml of a 0.005 to 0.1% solution of the polymer) by means of a mechanical inlet device. The pulse of the polymer solution injected into the column entry becomes diluted and attenuated as the different species are separated on the column packings. 

We now consider how the elution volume axis of a raw chromatogram, such as shown in Fig. 4.25, can be translated into a molecular weight scale. This necessitates a calibration of the particular GPC column for the particular polymer-solvent system used. Such a calibration requires the establishment of a relationship between the volume of solution eluted (or, equivalently, the elution time for a given flow rate of solution) and molecular weight of monodisperse fractions of the same polymer. The main problem encountered in this task is that monodisperse or very narrow distribution samples of most polymers are not generally available. However, such samples are available for a few specific polymers. A notable example is polystyrene for which anionically polymerized samples of narrow mole- 

Gel permeation chromatography (GPC) is a widely used technique for determining molar mass and molar mass distrihution of polymers. In its usual form it is not an absolute method, though hy making the appropriate measurements it may be made to be so. 

Gel permeation chromatograms actually give information about molecular size. For any polymer, size is determined hy a number of factors. These include not only molar mass but also temperature and thermodynamic quality of the solvent. Hence the relationship between size and molar mass is unique for each particular polymer-solvent combination, and we caimot assume that because two peaks of different polymers, even in the same solvent at the same temperature, have the same elution volume their molecules have the same molar mass. 

In practice, therefore, gel permeation chromatographs are usually cahbrated by running monodisperse specimens of known molar mass, and determining their elution volume. From the results, a graph is plotted of log M against 

Monodisperse polymers for this purpose may be obtained from commercial suppliers, but they are expensive and for many polymers not available in sufficiently narrow molar mass fractions. For this reason, the not very satisfactory procedure is sometimes adopted of calibration with polystyrene, since this polymer can be relatively easily prepared in discrete, monodisperse fractions using anionic polymerisation. The unknown polymer is then 

Yet as long ago as 1966 the problem of calibration in GPC was solved. In that year, Benoit and his co-workers recognised that GPC separates on the basis of the hydrodynamic volume of the polymer molecules in solution. The intrinsic viscosity [rj] is related to the hydrodynamic volume, V, by the equation  

Gel permeation chromatography (GPC), sometimes referred to as gel filtration chromatography (GFC) or size exclusion chromatography (SEC), entails the chromatographic fractionation of macromolecules according to molecular size. 

In later work, Forss et al. also analyzed kraft lignins by the same technique, using aqueous sodium hydroxide as the eluent (Forss et al. 1976, 1984). Fractionation of kraft lignins on Sephadex gels has also been carried out by other investigators (Connors 1978, Connors et al. 1978, 1980, Sarkanen et al. 1982, Yan et al. 1984, Kondo and McCarthy 1985, Dolk et al. 1986). 

Springer Series in Wood Science Methods in Lignin Chemistry (Edited by S.Y. Lin and C.W. Dence) 

In reality, the universal calibration ctine is not quite universal. Certain exceptions exist, such as polymers with very rigid backbones. However, the equation does apply to a very large number of systems, and this means that it is possible to calibrate the chromatograph with one set of readily available monodisperse standards and to use the restilts to determine the real molar mass of a different polymer. 

Calibration Flla - CAL8MAR (Cubic fit) Correction factor - Nona. 

The calibration curve can be divided into three sections. In the first, there is no separation of the small molecules as they are all totally occluded within the pores of the gel and there is no separation. The second has an approximately linear variation of the elution volume with molecular weight over a wide range of elution volume. The columns separate the polymer molecule over this particular range of molecular weights very well. In the final section of the 

IIUH w a Wtfl UI ihc differ turn dfebj file viIlm 

Since the detector response is proportional to the concentration of the eluant in the detector cell and independent of molecular weight, then, since the 

Gel permeation chromatography (GPC) involves the separation of molecules in solution based on their molecular size. The molecules to be separated, usually natural or synthetic polymers, are carried down a column of a porous gel with pores of varying sizes. Depending on the size of the polymer, it will pass into some of the pores. The more pores it passes into, the longer it will spend in the column. Thus the larger molecules are eluted first and the smaller last. GPC has little application in surfactant analysis apart from sample preparation where it is used to remove polymeric material from the components of interest prior to more detailed analysis. 

Gel permeation chromatography (GPC) is another important tool in the polyethylene characterization process. Although the Melt Flow Ratio (MFR) value obtained with an experimental sample of polyethylene 

1-hexene. Reprinted from Kirk-Othmer Encyclopedia of Chemical Technology, 

Three examples of the importance of GPC data in understanding the changes in polyethylene molecular structure due to modifications of ethylene polymerization catalysts will be discussed. 

This particular experimental catalyst produced a trimodal MWD with molecular weight ranging from 10 -10, while a modification of this 

Temperature rising elution fractionation (TREE) is especially useful in probing the branching content in low-density (LDPE) and linear low-density polyethylene (LLDPE). The concept of elution fractionation was described as early as 1950 when Desreux and Spiegel [18] deposited polyethylene sample onto a CeHte packed column and obtained the fractions by eluting the packed column with toluene at successively higher temperatures. 

The column is prepared and used as in ion-exchange chromatography. It is important, that the resin be. soaked with solvent for 2-48 hr before placing it in the column. 

Gels commonly used with polar substances (and aqueous solutions) are Sephadex,f Bio-Gel,t and Bio-BeadsJ. The last of these are available in a form usable with nonpolar substances and solvents. 

This technique has been used extensively by polymer chemists to characterize the molecular weight distribution of a polymer preparation and by biochemists to desalt or separate enzymes, nucleic adds, and polysaccharides. When the procedure is applicable, it can be used very effectively on a preparative scale. 

The technique of gel permeation chromatography (GPC) was developed during the mid-1960s, and is an extremely powerful method for determin- 

The volume of solvent contained in a GPC system from the point of solution injection, through the column to the point of concentration detection can be considered as the sum of a void volume Vq (i.e. the volume of solvent in the system outside the porous beads) and an internal volume Vi (i.e. the volume of solvent inside the beads). The volume of solvent required to elute a particular polymer species from the point of injection to the detector is known as its elution volume Ve and on the basis of separation by size-exclusion is given by 

For the flow rates typically used in GPC (about lcm min ) it is found that Ve is independent of flow rate. This means that there is sufflcient time for the molecules to diffuse into and out of the pores such that equilibrium concentrations, c,- and Cq, of the molecules are attained inside and outside the pores respectively. Since the separation process takes place under equilibrium conditions, Kse can be considered as tlie equilibrium constant 

For separation exclusively by size-exclusion, the enthalpy change associated with transfer of solute species into the pores must be zero. Thus AGp is controlled by the corresponding entropy change A5p and is given by AGp = -TASp. Hence from Equation (3.183), Kse is given by 

The number of conformations available to a polymer molecule is reduced inside a pore because of its close proximity to the impenetrable walls of the pore, and so A5 is negative. This loss of conformational entropy is equivalent to exclusion of the centre of mass of the molecule from regions closer to the walls of the pores than the hydrodynamic radius of the molecule (see Fig. 3.23). Methods of statistical thermodynamics can be used to obtain expressions for A5 on the basis of simplifying models (e.g. Gaussian coils entering cylindrical pores). The details will not be considered here, but the general result is of the form 

F re 5-1. Separation of large and small molecules using gel permeation chromatography. Molecules are represented schematically as large and small filled circles. 

Although comprehensive mathematical treatment of solute behavior on 

GPC is extremely valuable for both analytic and preparative work with a wide variety of systems ranging from low to very high molecular weights [2], The method can be applied to a wide variety of solvents and polymers, depending on the type of gel used. GPC has been used for routine polymer characterization and quality control, particularly in determinations of MWD and for characterizing low polymers and small molecules, e.g., for prepolymers in resins and for polymer additives [7,8]. 

The use of multidetector GPC greatly increases the power of SEC, particularly in the case of copolymers. For copolymers of styrene-maleic anhydride (SMA), not only can the molecular weight distribution be determined, using a differential refractive index (DRI) detector, but also the compositional information of SMA (styrene content or acid number) by combining chromatograms from DRI and UV detectors [9]. 

GPC is a further special form of liquid chromatography. The separation column is packed with porous, polymer gels (e.g. polystyrene gel) as stationary phase. The particle size of the packing material and the size distribution of the pores are well defined and uniform. In GPC molecules are separated according to their effective size in solution, i.e., their hydrodynamic volume, and not according to their affinity for the support material. 

Sample molecules that are too large to enter the pores of the support material, which is commercially available in various pore dimensions, are not retained and leave the column first. The required elution volume Ve is correspondingly small. Small molecules are retained most strongly because they can enter all the pores of the support material. Sample molecules of medium size can partly penetrate into the stationary phase and elute according to their depth of penetration into the pores (Fig. 7.3). No specific interactions should take place between the molecules of the dendrimer sample and the stationary phase in GPC since this can impair the efficiency of separation by the exclusion principal. After separation the eluate flows through a concentration-dependent detector (e.g. a UV/VIS detector) interfaced with a computer. One obtains a chromatogram which, to a first approximation, reflects the relative contents of molecules of molar mass M. If macromolecules of suitable molar mass and narrow molar mass distribution are available for calibration of the column, the relative GPC molar mass of the investigated dendrimer can be determined via the calibration function log(M) =f( Vc). 

GPC in dendrimer chemistry Since the principle of separation of GPC is based on the different sizes (hydrodynamic volumes) of the molecules, this is an ideal 

Macromolecules can be fractionated according to their constitution, configuration, or molecular weight by chromatographic methods. Adsorption chromatography is used rarely. Elution chromatography and gel-permeation chromatography are used much more often. 

The material to be separated is placed as a thin layer on an inert carrier and then eluted. Metal foil or quartz sand, for example, are suitable inert carriers. The metal foil is dipped into the macromolecular solution and then dried. The surface film is then eluted at constant temperature with solvent-precipitant mixtures of increasing solvent power. Thus, the lower molecular weight fractions are removed first. 

An elegant variant of this procedure is known as the Baker-Williams method. In this method, the chromatographic column is also surrounded by a thermostated heating jacket which ensures that an adequate thermal gradient is maintained. The separation efficiency is enhanced by the simultaneous concentration and temperature gradients. 

In gel-permeation chromatography, the separating column consists of a solvent-swollen gel with pores of various diameters. An 0.5% solution 

9 Dererminatim) of Molecular Weight. Molecular-Weight Distrihulion 

It is now generally accepted that the GPC retention volume is a function of the product M tf, independent of the nature or structure of the polymer 46, 47) though Pannell 45) found that it failed to correlate the elution behaviour of his highly branched polystyrenes, it may be accepted that M rf will be determinable from GPC retention volumes for moderately branched polymers. To estimate branching, it is necessary to separate this product so that M and [rf are both known and the relation between them can then be used, subject to the uncertainties mentioned in Subsection 9.2.2, for this purpose. It is usual to measure rf rather than M in order to make the separation, as it is easier. The combination of GPC and intrinsic viscosity measurements is now the most usual method for studying long branching. 

Tung (106) has proposed the combination of sedimentation constant data with those from GPC, so as to obtain a distribution of branching, without fractionation being required. 

After passage through the column(s), the solvent stream (eluant) carrying the size-separated polymer molecules passes through a detector, which responds to the weight concentration of polymer in the eluant. The most commonly used detector is a differential reffactometer. It measures the difference in refractive index between the eluted solution and the pure solvent. This difference is proportional to the amount of polymer in solution. Spectrophotometers are also used as alternative or auxiliary detectors. 

The elution volume (also called the retention volume) is the volume of solvent that has passed through the GPC column from the time of injection of the sample. It is conveniently monitored by means of a small siphon, which actuates a marker every time it fills with eluant and dumps its contents. The raw GPC data are thus available as a trace of detector response proportional to the amount of polymer in solution and the corresponding elution volumes. A typical GPC record (gel permeation chromatogram) is shown in Fig. 4.19. 

The molar mass associated with the elution volume is probably represented by the following average  

This average gives, for a Schulz-Flory distribution of degree of coupling k. 

Consequently, Mgpc ) = Mw ) for spheres, where ar, = 0, and Mgpc ) = Afz +1 for rigid rods. An average close to the weight average is obtained for coil-shaped molecules with the usual distributions when the experiments are run in theta solutions. 

The width of the elution curve increases with the width of the distribution. But unimolecular substances do not give a sharp signal an elution curve is also obtained. This is produced by what is known as the axial dispersion. According to the model described above, it signifies a distribution of residence times in the pores. The effect must also be taken into account when calculating molar mass distributions. For this, it is assumed that the total standard deviation atm is composed of the standard deviation associated with molecular inhomogeneity Omoi and the standard deviation resulting from axial dispersion Oad  

Oad is determined by reversing the flow of the liquid after development of the elution curve. The amoi can be calculated, and the true elution curve can be subsequently constructed. 

This technique has found limited applications in polymer additive analysis in plastics and rubbers. These include aromatic amines and antiozonants in rubber extracts [2, 50-62], and di-laurylthiodipropionate in polymer extracts [56]. 

Protivova and Pospisil [19] have reported on the behaviour of some amine antioxidants and antiozonants and some model substances (phenols, aromatic hydrocarbons and amines) 

Haulein, Corning Glass Works, Personal Communication, 1970. 

Crompton, Polymer Reference Book, Rapra Technology, Shrewsbury, UK, 2006. 

Chemical Derivatization in Analytical Chemistry, Volume 1, Eds., R.W Frei and J.F. Lawrence, Plenum Press, New York, NY, USA, 1981. 

While use of the viscosity-average molecular weight of a polymer in calibrating K and a in equation (3.97), My values usually are not initially known. The calibration problem may be alleviated by one or more of the following methods  

Use of polymers prepared with narrow molecular weight distributions, such as via anionic polymerization. 

Use of weight-average molecular weight of the polymer, since it is closer than the number-average molecular weight. 

Say we are interested in a fast, approximate molecular weight of a polystyrene sample. We dissolve 0.10 g of the polymer in 100 ml of butanone and measure the flow times at 25°C in an Ubbelhode capillary viscometer. The results are 

Starting with equation (3.84), and nothing that the flow time is proportional 

This technique has been applied to the determination of organic compormds and cations induding crude oils, fulvic adds, tarmins, carbohydrates, trihalomethane precursors and lithium. 

The technique separates components of a mixtiure in order of their molecular size, pradionally their molecular weight. In a typical example [141, a sample of a crude oil was dissolved in toluene or tetrahydrofuran and pumped flwough a column of 60A Styragel. The effluent was monitored by a differential refractometer. In the case of cations, separations based on molecular size have been achieved with alkali metals eluting in order potassium, sodium, lithium, magnesium and caldum 

The calibration curve is best prepared using monodisperse samples of the same polymer in the same solvent and at the same temperature. Unfortunately, 

Thomas, D. R, and R. S. Hagan, The Influence of Molecular Weight Distribution on Melt Viscosity, Melt Elasticity, Processing Behavior, and Properties of Polystyrene, Polym. Eng. Set, 9, 164-171, 1969. 

Polymer Molecular Weight Methods, Advances in Chemistry Series No. 125, American Chemical Society, Washington, DC, 1973. 

Taylor, Determination of Molecular Weight of Nylon, Anal. Chem., 19, 448-451, 1947. 

Orofino, Nylon 66 Polymers I. Molecular Weight and Compositional Distribution, J. Polym. Set, A2(7), 1-25, 1969. 

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Gel permeation chromatography, exclusion chromatography. gel filtration chromatography. A technique for separating the components of a mixture according to molecular volume differences. A porous solid phase (a polymer, molecular sieve) is used which can physically entrap small molecules in the pores whilst large molecules pass down the column more rapidly. A solvent pressure up to 1000 psi may be used. 

Two classes of micron-sized stationary phases have been encountered in this section silica particles and cross-linked polymer resin beads. Both materials are porous, with pore sizes ranging from approximately 50 to 4000 A for silica particles and from 50 to 1,000,000 A for divinylbenzene cross-linked polystyrene resins. In size-exclusion chromatography, also called molecular-exclusion or gel-permeation chromatography, separation is based on the solute s ability to enter into the pores of the column packing. Smaller solutes spend proportionally more time within the pores and, consequently, take longer to elute from the column. 

At first glance, the contents of Chap. 9 read like a catchall for unrelated topics. In it we examine the intrinsic viscosity of polymer solutions, the diffusion coefficient, the sedimentation coefficient, sedimentation equilibrium, and gel permeation chromatography. While all of these techniques can be related in one way or another to the molecular weight of the polymer, the more fundamental unifying principle which connects these topics is their common dependence on the spatial extension of the molecules. The radius of gyration is the parameter of interest in this context, and the intrinsic viscosity in particular can be interpreted to give a value for this important quantity. The experimental techniques discussed in Chap. 9 have been used extensively in the study of biopolymers. 

This chapter contains one of the more diverse assortments of topics of any chapter in the volume. In it we discuss the viscosity of polymer solutions, especially the intrinsic viscosity the diffusion and sedimentation behavior of polymers, including the equilibrium between the two and the analysis of polymers by gel permeation chromatography (GPC). At first glance these seem to be rather unrelated topics, but features they all share are a dependence on the spatial extension of the molecules in solution and applicability to molecular weight determination. 

Gel permeation chromatography (GPC) is the term found most widely in the polymer literature. In this context it is used most widely as an analytical... 

Salt Effects. The definition of a capacity factor k in hydrophobic interaction chromatography is analogous to the distribution coefficient, in gel permeation chromatography ... 

Bentone-34 has commonly been used in packed columns (138—139). The retention indices of many benzene homologues on squalane have been determined (140). Gas chromatography of C —aromatic compounds using a Ucon B550X-coated capillary column is discussed in Reference 141. A variety of other separation media have also been used, including phthaUc acids (142), Hquid crystals (143), and Werner complexes (144). Gel permeation chromatography of alkylbenzenes and the separation of the Cg aromatics treated with zeofltes ate described in References 145—148. 

MFI = melt flow index IV = intrinsic viscosity in CH2CI2 at 25°C From gel-permeation chromatography using polystyrene standards. 

Fig. 3. Molecular weight distribution curves as determined by gel-permeation chromatography. A represents i9f2v (9-phthahc resins B, highest molecular...

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