Preparative Fractionation

Larger injection volumes, e.g., 2% of the total column volume, are sometimes advantageous in the preparative fractionation of polymers (33). More samples can be injected using larger injection volumes with a slight decrease in resolution. When the same amount of sample is injected with a smaller injection volume and a higher sample concentration, the resolution decreases more significantly. 

The latest trend is to smaller beads in smaller columns, as this saves eluent and shortens the time for a chromatographic analysis. This argument can be correct if only one suitable detector is used. However, these modern small columns are not optimal for a combination of detectors. So-called multiple detection is a combination of some detectors with different measurement principles (differential refractometer, spectral photometer, light-scattering detector, on-line viscometer) behind the last column, mostly in series, seldom in a branched ( parallel ) order. In this way, the tedious preparative fractionation of a polymer sample can often be avoided. 

The next approach was to reduce the polydispersity of the chain length by fractionation. In the earliest preparation, fractionation was performed by precipitation or by dialysis, but the results were not very satisfying. 

Preoperative mechanical bowel preparation fractional doses of sodium phosphate on days 3 and 2 prior to surgery... 

It is often necessary to assess the efficiency of cell fractionation procedures. Electron microscopy of the prepared fractions is very informative but gives no quantitative indication of the purity of the fraction. It is often easier to measure the relative concentrations of marker enzymes in each fraction (Table 8.9). 

The migration order of wine anthocyanins in CE has been studied in detail and the results have been compared with those obtained by RP-HPLC-MS. Wines were filtered and used for the analyses without any other pretreatment. Wine samples of 10 ml were freeze-dried, redissolved in methanol and applied for semi-preparative fractionation. CZE measurements were carried out in a fused-silica capillary (46 cm effective length, 75 /an i.d.). The capillary was conditioned with 0.1 M NaOH (2 min), water (2 min) and running buffer (5 min). The buffer consisted of 50 mM sodium teraborate (pH = 8.4) containing 15 per cent (v/v)... 

Three other components that my laboratory has identified and partially purified from Fraction 2 of reticulocytes, termed CF1-CF3, are involved in the degradation of proteins ligated to ubiquitin [24]. These are apparently subcomplexes of the 26S proteasome, a large ATP-dependent protease complex first described by Re-chsteiner and co-workers [25], CF3 is identical to the 20S proteasome core particle [26], while CFl and CF2 may be similar to the base and lid subcomplexes of the 19S regulatory particle of the 26S proteasome, described more recently by the Finley laboratory [27], In hindsight, the reason for finding subcomplexes, rather than the complete 26S complex in Fraction 2 was technical we have routinely prepared Fraction 2 from ATP-depleted reticulocytes [20], under which conditions the 26S proteasome dissociates to its subcomplexes. We found that incubation of the three subcomplexes in the presence of ATP promotes their assembly to the 26S proteasome [24, 26]. The role of ATP in the assembly of the 26S proteasome complex remains unknown. 

Preparative fractionation of lOOmg sample of HPD and the product of the DP-HP synthesis were carried out on a 5x30cm Sephadex LH-20 column (Pharmacia, Piscataway, NJ.). The porphyrin components were first dissolved in 40ml of tetrahydrofuran-methanol-5mM aqueous phosphate buffer, pH 7, (2 1 1), and subsequently eluted with the same solvent. The use of this methodology for the isolation of the tumor-localizing fraction of HPD from other porphyrin components present in the drug mixture has been discussed in detail in the literature.(11) The component of HPD which elutes very close to the void volume of the LH-20 column has been characterized as the tumor-localizing fraction of HPD. 

Partly purified preparation fractionated from T. reesei culture filtrate by gel chromatography (10). 

As explained in Sections 16.3.4, 6.4.1, and 16.4.2, SEC is a nonabsolute method, which needs calibration. The most popular calibration materials are narrow molar mass distribution polystyrenes (PS). Their molar mass averages are determined by the classical absolute methods—or by SEC applying either the absolute detection or the previously calibrated equipment. The latter approach may bring about the transfer and even the augmentation of errors. Therefore, it is recommended to apply exclusively the certified well-characterized materials for calibrations. These are often called PS calibration standards and are readily available from numerous companies in the molar mass range from about 600 to over 30,000,000g moL. Their prices are reasonable and on average (much) lower than the cost of other narrow MMD polymers. Other available homopolymer calibration materials include various poly(acrylate)s and poly(methacrylate)s. They are, similar to PS, synthesized by anionic polymerization. Some calibration materials are prepared by the methods of preparative fractionation, for example, poly(isobutylene)s and poly(vinylchloride)s. 

Fractionation according to molecular size ( molar mass ) The components of a sample are separated according to their molecular size caused be their ability to penetrate the macoporous beads. Examples Analytical or preparative fractionation of macromolecules, separation of oligomers from monomers. 

Tetrahydrofuran (THF, UV grade) was used as the mobile phase throughout this work since the extent of fractionation could be demonstrated by direct analysis of the preparative fractions without the need for concentration. When samples are to be recovered by removal of solvent, other mobile phases (methylene chloride, etc.) may be preferred to avoid concentrating solvent impurities which are formed in THF on exposure to air unless additional precautions are taken. 

Figure 11. GPC analysis of epoxy starting material and preparative fractions.

A Waters Associates Anaprep GPC fitted with one 4 ft X 2.4 inches od Styragel column having a nominal porosity of 104 A was used for the preparative fractionation of the PMMA blend in tetrahydrofuran at a temperature of 25 °C and at a flow rate of 30 ml/min. The degasser and differential refractometer were operated at 35° and 25 °C, respectively. Samples having concentrations of 0.25 wt-vol % were respectively, automatically injected from a 100 ml loop over a 5-minute period. Ten 125 ml fractions were automatically collected for each sample injection. Upon... 

Preparative fractionation of PS with pyrene or OH endgroups was conducted on a silica column (350 x50 mm d0 = 6nm dP 100 pm). The eluent was benzene-cyclohexane (70 30) or chloroform with stepwise increased methanol content (0.01 to 2 %). The fractions obtained formed a series decreasing in molar mass. This might have been due to size exclusion although the column possessed adsorption activity. 

Egeberg, P. K., and Alberts, J. J. (2003). HPSEC as a preparative fractionation technique for studies of natural organic matter (NOM). Environ. Technol. 24,309-318. 

Figure 8. Enantiomeric purity of the isolated invertomers. Left racemic sample middle pure (R)-enantiomer right (SJ-enantiomer contaminated with (R)-enantiomer from incomplete preparative fractionation (ee = 98 %). Analytical column 25 m x 0.25 mm glass capillary coated with 0.125 molal nickel(II) bis[(2-heptafluorobutanoyl)-(TS,5S/)-4-methylthujonate] (Ni METHU2) in OV-101 at 50°C. The (R)-enantiomer is eluted before the (S)-enantiomer on (IS,5S)-Ni-METHU2 (Schurig and Leyrer, 1990).

The objective of this communication is to report structural information on a bituminous coal from a thorough characterization of its extracts. The characterization was carried out according to a scheme devised on the basis of our current understanding of coal chemistry. The scheme consists of considerations in preparation, fractionation, and analysis of coal-derived liquids (CDL) to obtain molecular-level information on the CDL per se as well as on the parent coal. The information obtained relates mostly to the structure of component clusters. 

Applying FFF techniques for preparative separations in the form of SPLITT channels also seems to be very promising. Here, future developments must concentrate not only on the application of other physical fields for the separation in SPLITT but also on the coupling of various SPLITT channels with different cut off limits to design a preparative fractionator for broadly distributed samples. 

The first preparative fractionation of betalaine pigments by means of ion-pair high-speed conntercurrent chromatography utilized a solvent system composed of trifluoroacetic acid at low concentration as the IPR that considerably improved affinity of polar betacyanins and betaxanthins to the organic stationary phase of the biphasic solvent mixture [19],... 

Used also to prepare fractions for subsequent additional separations and/or instrumental analysis. 

With a chromatographic technique capable of routinely yielding preparative fractions, quantitative and C FT NMR was the major spectroscopic tool used for chemical characterization. The established utility of and C NMR for characterization of coal products is documented well. Unfortunately, high-resolution C FT NMR is not quantitative normally under operating conditions used typically. (It should be noted that quantitative FT NMR measurements also are not obtained routinely. The problem of variable spin lattice relaxation times (Ti s) is present also in FT NMR. In addition, the greater signal intensity of NMR in comparison with C FT NMR poses an additional potential problem of detector linearity in the FT NMR receiver.) For C FT NMR, variable spin lattice relaxation times (Ti s) and nuclear Over-hauser effects (a result of pseudo random noise decoupling) usually... 

Jl ield-flow fractionation (FFF) is a separation method convenient for the analysis and characterization of macromolecules and particles of synthetic or natural origin. Under the appropriate experimental conditions, it also can be applied for the preparative fractionation. The separation is due to a simultaneous action of the effective field forces and of the carrier liquid flow inside an open channel on the dissolved or suspended macromolecules or particles. The carrier liquid flows in the direction of the channel longitudinal axis and the field forces act in the perpendicular direction across the channel thickness. Each component of the fractionated sample interacting with the field forces is selectively transported across the channel. This concentrating process... 

The focusing FFF can be used for continuous preparative fractionation 7,8). If the fractionation channel is equipped with several outlet capillaries at various positions in the direction of focusing and the sample to be fractionated is continuously pumped into the channel, the focused layers eluting through the individual outlets can be collected. 

Altria, K.D. Dave, Y.K. Peak homogeneity determination and micro-preparative fraction collection by capillary... 

There can be little doubt that, during most preparative, fractionation methods, a certain amount of degradation is inadvertently introduced. As a result, the intrinsic viscosities of both fractions of starch will be found to be lower than the corresponding values for their native state. In order to make a comparison possible, some method of fractionation has to be developed which gives no degradation whatsoever. In this respect, the techniques outlined in this Section might all have a fair chance of success. Restricting attention to potato starch as a substrate, and furthermore to the intrinsic viscosity of its amylose fraction as measured in 1.0 AT potassium... 

While ultrafiltration has been used extensively by water chemists in the isolation of humic substances (Schnitzer, 1978), its use with soil materials has been more limited (Cameron et al., 1972b Wake and Posner, 1967). From the foregoing discussion it should be clear that, as a technique for preparative fractionation and desalting, ultrafiltration is very attractive and will rival gel permeation chromatography in this type of work. Only time and a considerable amount of effort will tell which is the superior technique. 

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