Micron Spherical Silica Gel Powder for Cosmetics Additive and Column Chromatography
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Review of Practical Application of Silica Gel in Column Chromatography(I)

Chromatography, also known as chromatography. It is a distribution method based on distribution equilibrium as the mechanism. The chromatography system contains

two phases, one is the stationary phase and the other is the mobile phase. When the two phases move relative to each other, the differences in the distribution equilibrium

properties of each component contained in the mixture are repeatedly utilized to achieve the purpose of separation from each other. It is a method of purifying and

separating organic or inorganic substances.

Chromatography can be divided into three types according to the state of the stationary phase: column chromatography, plate chromatography and rod chromatography.

Column chromatography silica gel and thin-layer silica gel plates are commonly used in laboratories and practice, as well as their coordinated applications.

Column chromatography silica gel

1. Principle of adsorption chromatography

Under certain conditions, there is an interaction between silica gel and the separated substance. This interaction is mainly physical and chemical. The two physical effects come from the van der Waals force between the surface of the silica gel and the solute molecules. The chemical effect is mainly the hydrogen bonding between the silanol groups on the surface of the silica gel and the substances to be separated.

2. Practical application of column chromatography silica gel

A glass chromatography column, an iron stand, a beaker, an Erlenmeyer flask, a glass funnel with a larger diameter, and a glass rod.

2.3 Packing the column

2.3.1 Addition of silica gel to column chromatography

There are two methods: dry method and wet method. The operation method is introduced in detail below.

① Dry method: Add silica gel to the chromatographic column at once, vibrate the tube wall to make it sink evenly, and then slowly add the mobile phase used when starting chromatography along the tube wall, or add a piston to the outlet at the lower end of the chromatographic column and add an appropriate amount of mobile phase. , unscrew the piston to allow the mobile phase to slowly drip out, and then slowly add silica gel from the top of the tube to make it evenly wet and sink, forming an adsorption layer with moderate tightness in the tube. During the operation, sufficient mobile phase should be kept on top of the silica gel.

② Wet method: Mix the silica gel with the mobile phase, stir to remove air bubbles, slowly pour it into the chromatographic column, and then add the mobile phase to wash off the adsorbent attached to the tube wall to make the surface of the chromatographic column smooth. The mobile phase used to fill the silica gel naturally flows down from the chromatographic column. When the liquid level is equal to the surface of the column, the sample solution is added.

2.3.2 Loading samples

Dissolve the sample in the mobile phase used in chromatography, and then slowly add it along the wall of the chromatography column. Be careful not to flip up the silicone. Or dissolve the sample in an appropriate solvent. Mix with a small amount of silica gel, and then evaporate the solvent to make it loose; add the silica gel mixed with the sample on the prepared chromatographic column. If the sample is insoluble in a common solvent, grind the sample and an appropriate amount of silica gel in a mortar and mix evenly before adding.

2.4 Elution and collection

Unless otherwise specified, the type and proportion of the mobile phase are usually changed incrementally according to the elution capacity of the mobile phase, and the effluent is collected separately until the components contained in the effluent are significantly reduced or no longer contained, then change the type and proportion of mobile phase. During the operation, sufficient mobile phase should be kept on top of the silica gel.

2.5 Detection after sample separation

2.5.1 Preliminary detection

When a certain amount of flushing solvent flows out, the effluent can be preliminarily tested, and the Erlenmeyer flask is replaced with a small test tube for collection.

Generally, only a preliminary quick test is performed, so a thin layer plate is usually taken, use a pencil and a ruler to divide the silicone plate into small squares, and number them in a certain order. Take a glass capillary tube with an inner diameter of about

0.3mm, dip it in a small amount of the effluent, and point it in a small grid of the thin layer plate. After it is half dry, use physical or chemical methods to detect it.

2.5.2 Formal testing

①Spotting: Take the flushing solutions collected separately and spot them directly. If the flushing solution is too dilute and the concentration is too small, it can be

concentrated first. The container for spotting usually uses a glass capillary tube, and the diameter of the spot is generally 3-5mm.

② Expansion: It is carried out in an ordinary expansion slot, and the expansion method is often upward expansion.

③Developing agent: practical flushing solution.

④ Color development: Generally, physical detection methods and chemical detection methods are commonly used. Among the physical detection methods, there is first

the ultraviolet light method. There are two commonly used wavelengths of ultraviolet light (254nm and 365nm), followed by the iodine vapor colorimetric method.

Chemical detection methods usually use direct spraying of color developers. The color developers include general color developers and special color developers. The most

common general color developers are sulfuric acid-ethanol or methanol (1:1) solution, after spraying, some compounds react immediately, but most compounds need to be heated for several minutes to develop color. Different compounds react differently,

so the colors are often different. Special chromogenic reagent refers to a reagent that develops color for a certain or a certain type of compound. It uses the unique

properties of the compound itself or uses the special reaction of certain functional groups it contains to develop the color according to the spots.

The content of the substance to be separated can be roughly estimated.

2.6 Merge processing

According to the results of the above thin layer plate detection, we can combine the parts with the same Rf value, and then use a rotary evaporator to rotary evaporate

the combined parts to obtain the target product we need.

2.7 Column washing

In most cases, the silica gel in the chromatography column is used once, but after use, the chromatographic column also contains flushing solvent.Therefore, it is difficult to remove the silica gel inside. One way to remove the silica gel is to leave the column tube for a period of time, let the solvent evaporate naturally,

and then pour out the silica gel. But this method is time-consuming and environmentally polluting. The second method can be to use a wooden or plastic rod slightly

longer than the chromatographic column to take out the silica gel containing the solvent segment by segment, but this method is also more troublesome. The third

method is to use a general vacuum pump to extract the remaining solvent in the chromatographic column under reduced pressure, and add a cold trap between the

chromatographic column and the vacuum pump. In this way, the extracted solvent is effectively collected without polluting the environment. The silica gel in the

chromatographic column can be dried quickly, so that the silica gel can be poured out easily.