1. Structure of liquid chromatography column:
a. The empty column is assembled from column joints, column tubes and filters.
The column joints are constructed with a low dead volume. The column joints are threaded assemblies at both ends, with 7/16-inch external threads on one end and 3/16-inch internal threads on the other end (standardized at home and abroad). The 7/16-inch external thread is connected to the 1/4-inch column tube (Φ6.35mm) and placed in the middle for sealing. The 3/16" internal thread is connected to the 1/16" (Φ 1.57mm) connecting tube, and a pressure ring is placed in the middle for the seal of the column joint. In order to minimize the dead volume outside the column, when installing the column, the Φ1.57mm connecting tube should be inserted into the bottom by the hollow screw, and then the hollow screw should be tightened. The pressure ring is pressed and deformed by the hollow screw and then hung on the connecting pipe (the length of the pipe exposed after the connecting pipe passes through the pressure ring should be strictly controlled to 2.5 mm long or other fixed size).
In the column joints at both ends, a stainless steel filter (or strainer) is placed at each end of the column tube to block the column packing from being washed out of the column by the mobile phase. Each component of the empty column is made of 316# stainless steel, which can withstand the general solvent effect. However, due to the corrosive nature of the chloride-containing solvent, it should be noted that the solvent and the connecting tube cannot be stored for a long time to avoid corrosion.
b, column packing:
The separation of the liquid chromatography column is carried out between the filler and the mobile phase, and the classification of the column depends on the type of filler.
Normal phase column: mostly silica gel as column packing. According to the appearance, it can be divided into two types: amorphous and spherical, and its particle diameter is in the range of 3-10 μm. Another type of normal phase filler is silica gel surface bonding - a functional group such as CN, -NH2, so-called bonded phase silica gel.
Reversed-phase column: A non-polar filler that is mainly based on silica gel and has an octadecyl functional group (ODS) bonded to its surface. There are also amorphous and spherical types.
Other commonly used reversed-phase fillers are also bonded to C8, C4, C2, phenyl, etc., and the particle size is between 3 and 10 μm.
2. Installation of the column:
a. Unpack the column and confirm the type, size, date of manufacture, and solvent stored in the column.
b. Unscrew the sealing plugs at the ends of the column and put them back in the box for spare.
c. According to the flow direction indicated on the column tube, connect the inlet end of the column to the inlet of the injection valve through the connecting pipe (if the conditions permit, it is recommended to use the guard column in front of the column); the outlet of the column is connected to the detector. The connecting pipe is a stainless steel pipe having an outer diameter of 1.57 mm and an inner diameter of 0.1 to 0.3 mm. Both ends of the connecting tube have a hollow screw and a sealing ring for sealing. When taking over, be sure to try to reduce the dead volume outside the column. After connecting the tube through the hollow screw and the pressure ring, insert it as far as possible into the end, then tighten the cannulated screw clockwise until it stops, then use the wrench to continue to screw 1/4-1/2 turn clockwise, remember not to use too much force. If the column is leaking after being pressurized by the mobile phase, use a wrench to continue to turn 1/4 turn clockwise until it does not leak.
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