Gel Exclusion chromatography.pptx

Introduction, methodology and applications are discussed

1. Gel Exclusion Chromatography By Dr. Nidhi Gupta Assistant Professor M.M. College of Pharmacy Maharishi Markandeshwar (Deemed to be University), Mullana , Ambala , India

2. Introduction Gel permeation chromatography (GPC) is a form of size exclusion chromatography (SEC) that uses organic solvents to separate analytes based on their size. Polymer analysis is a popular application of this technique. SEC was first developed as a technique by Lathe and Ruthven in 1955. The term gel permeation chromatography was coined by J.C. Moore of the Dow Chemical Company, who researched the technique in 1964 and licensed the patented column technology to Waters Corporation, which commercialized it in 1964.

3. Principle of Gel Exclusion chromatography The analytes are separated by GPC based on their size or hydrodynamic volume (radius of gyration). Other separation methods, on the other hand, rely on chemical or physical interactions to distinguish analytes . Porous beads packed in a column are used to separate the particles. Since smaller analytes can penetrate pores more quickly, they spend more time in them and hence have a longer retention time. Since these smaller molecules spend more time in the column, they elute later. Larger analytes , on the other hand, spend little or no time in the pores and are easily eluted. A number of molecular weights can be divided in each column .

4. Analytes that are too large will not be retained; on the other hand, analytes that are too small will be entirely retained. Analytes that are not retained are eluted with the free volume outside of the particles (Vo), while those that are completely retained are eluted with the volume of solvent held in the pores. The following equation can be used to calculate the total volume, where Vg is the volume of the polymer gel and Vt is the total volume: Vt = Vg + Vi + Vo

5. Methods of Gel Filtration Chromatography Almost all gel permeation chromatography is done in chromatography columns. The experimental model is very similar to that of other liquid chromatography techniques. Samples are dissolved in a suitable solvent, which in the case of GPC is usually organic, and then filtered before being injected onto a column. The column is where a multi-component mixture is separated. The use of a pump ensures a steady supply of fresh eluent to the column. A detector is needed because most analytes are not visible to the naked eye. To obtain additional information about the polymer sample, several detectors are often used. The stationary process for GPC is gel. In order to apply a gel to a specific separation, the pore size of the gel must be carefully monitored. The absence of ionizing groups and low affinity for the substances to be separated in a given solvent are also desirable properties of the gel-forming agent.

6. Microporous packing material is used to fill the GPC column. The gel is poured into the column known as the gel filtration column. The eluent (mobile phase) should be a good solvent for the polymer, allowing the polymer to have a high detector response and wetting the packing surface. Tetrahydrofuran is the most common element in GPC polymers that dissolve at room temperature (THF). Piston or peristaltic pumps are the two types of pumps available for uniform distribution of relatively small liquid volumes for GPC. In GPC, a detector will continuously monitor the polymer concentration by weight in the eluting solvent. There are several different types of detectors, which can be classified into two groups. UV absorption, differential refractometer (DRI) or refractive index (RI) detectors, infrared (IR) absorption, and density detectors are the first types of concentration-sensitive detectors.

7. Application of Gel Filtration Chromatography GPC is often used to determine the relative molecular weight of polymer samples as well as molecular weight distribution. The molecular volume and shape function, as determined by the intrinsic viscosity, are what GPC truly measures. This relative data can be used to calculate molecular weights within 5% precision if comparable criteria are used. To calibrate the GPC, polystyrene standards with disparities of less than 1.2 are commonly used.

8. Disadvantages of Gel Permeation chromatography GPC, on the other hand, has several drawbacks. First, the number of peaks that can be resolved within the short time frame of a GPC run is small. Furthermore, for a satisfactory resolution of peaks, GPC as a technique needs at least a 10% difference in molecular weight. When it comes to polymers, the molecular masses of most of the chains are too close together for the GPC separation to produce anything other than large peaks. Another downside of GPC for polymers is that it needs filtration prior to use to prevent dust and other particulates from destroying the columns and interfering with the detectors.

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