Size exclusion chromatography, HPLC

High Performance Size Exclusion Chromatography (HPLC-SEC)... 

PEG amine was prepared from the bromide by Johansson s method, eq. 6, and by reductive amination of PEG aldehyde, eq. 7. Size exclusion chromatography (HPLC-SEC) revealed no noticable alteration in molecular weight distribution. 

Size-exclusion chromatography can be carried out using conventional HPLC instrumentation, replacing the HPLC column with an appropriate size-exclusion column. A UV/Vis detector is the most common means for obtaining the chromatogram. 

Showa Denko K.K. started the Shodex HPLC business in 1973 by developing columns to determine the molecular weight distributions of polymers produced at its petrochemical plant. Since then, more than 600 items of columns have been developed to achieve various kinds of analyses. Among them are several series of columns that can be used for size exclusion chromatography. The abundant variety of columns is one of the important characteristics of Shodex. Any kind of analytical requirements can be satisfied by choosing the appropriate column supplied by Showa Denko. 

Other groups have also used EC and CE to perform non-comprehensive multidimensional separations (15, 16). A three-dimensional separation was performed by Stromqvist in 1994, where size exclusion chromatography (SEC), reverse-phase HPLC, and CZE were used in an off-line manner to separate peptides (17). The most useful information gained from all of these non-comprehensive studies was knowledge of the orthogonality and compatibility of EC and CE. 

Stopher and Gage used size-exclusion chromatography (SEL) coupled to reversed phase HPLC for the direct injection of plasma in the analysis of an antifungal agent, voriconazole (12). Their system consisted of three columns, i.e. first a size-exclusion... 

Figure 3 Reversed-phase chromatography of products after alkaline hydrolysis of /3-poly(L-malate), Discrete polymer products are formed, which differ in length by several units of L-malate. The absorbance at 220-nm wavelength was measured, (a) /3-Poly(L-malate) before hydrolysis, (b) After 10-min incubation in 20 mM NaOH at 37°C. (c) After 15 h in 20 mM NaOH at 37°C. (d) After I h in 500 mM NaOH at 100°C. High pressure chromatography (HPLC) on Waters reversed-phase Ci8- i-Bondapak. The methanol gradient (in water-trifluoro acetic acid, pH 3.0) was programmed as follows 0-40 min 0.3-23%, 40-47 min 23-40%, 47-49 min 40%, 49-54 min 40-0%. (d) Inset size exclusion chromatography after 3-min alkaline hydrolysis at pH 10.2. BioSil SEC 250 column of 300 mm x 7.8 mm size, 0.2 M potassium phosphate buffer pH 7.0.

A number of analytical techniques such as FTIR spectroscopy,65-66 13C NMR,67,68 solid-state 13 C NMR,69 GPC or size exclusion chromatography (SEC),67-72 HPLC,73 mass spectrometric analysis,74 differential scanning calorimetry (DSC),67 75 76 and dynamic mechanical analysis (DMA)77 78 have been utilized to characterize resole syntheses and crosslinking reactions. Packed-column supercritical fluid chromatography with a negative-ion atmospheric pressure chemical ionization mass spectrometric detector has also been used to separate and characterize resoles resins.79 This section provides some examples of how these techniques are used in practical applications. 

Advances in size-exclusion chromatography, coupled with refractive index, absorption, viscosity, and lightscattering detectors, and MALDI-ToFMS, have made it possible to accurately determine molecular weight distribution (oligomer profiling), even at the relatively low values of polymeric additives (up to about 5000 Da). Advances in column design, e.g. high-resolution PS/DVB columns (> 105 plates m-1) mean that SEC can provide a valuable alternative to conventional HPLC techniques for the separation of small molecules. 

Detection in 2DLC is the same as encountered in one-dimensional HPLC. A variety of detectors are presented in Table 5.2. The choice of detector is dependent on the molecule being detected, the problem being solved, and the separation mode used for the second dimension. If MS detection is utilized, then volatile buffers are typically used in the second-dimension separation. Ultraviolet detection is used for peptides, proteins, and any molecules that contain an appropriate chromophore. Evaporative light scattering detection has become popular for the analysis of polymers and surfactants that do not contain UV chromophores. Refractive index (RI) detection is generally used with size exclusion chromatography for the analysis of polymers. 

Aurora Biomolecules dedicates to peptide synthesis (and polyclonal antibody production) for any small quantity purpose. FMOC chemistry (on Perceptive Biosystems Pioneer instruments) is used for peptides synthesis Online monitoring of the coupling efficiencies and HATU activation helps insure that the major component of the synthesis is the correct oligopeptide. Purification is firstly carried out by size exclusion chromatography, and then by HPLC on a PE vision purification workstation. Typically, 20 mg of pure peptide are obtained. The molecular weight of the purified peptide is determined as a final confirmation of quality. 

Analytical size exclusion chromatography was performed on a Waters HPLC system using a Biosil SEC-400 column (Biorad) acc. to [Kuhl et at. 2000] in order to examine the oligomerization status ofPSl and PS2. 

The difference in movement rates of various compounds through a column is attributed to differential migration in HPLC. This can be related to the equilibrium distribution of different compounds such as X, Y, and Z between the stationary phase and the flowing solvent(s), or mobile phase. The speed with which each compound moves through the column (ux) is determined by the number of molecules of that compound in the moving phase, at any moment, since sample molecules do not move through the column while they are in the stationary phase. The molecules of the solvent or mobile phase move at the fastest possible rate except in size exclusion chromatography, where molecular... 

ASE using dichloromethane has been applied to extract alkylphenols and short-chain NPEO from sediment [8,47]. Samples of 2-5 g were extracted in two cycles of 30 mL at 100°C and 69 atm. Clean-up was performed using size exclusion chromatography to remove high molecular weight lipids, and then using normal phase HPLC. 

Although the overwhelming majority of theoretical papers in liquid chromatography are dealing with the various aspects of RP-HPLC separation, theoretical advances have also been achieved in some other separation modes. Thus, a theoretical study on the relation between the kinetic and equilibrium quantities in size-exclusion chromatography has been published, hi adsorption chromatography the probability of adsorbing an analyte molecule in the mobile phase exactly r-times is described by... 

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