How to Choose the Right HPLC Column
Today we will take about how to choose the right column. The HPLC column to be simple frankly it was made by a stainless tube and packing material (stationary phase).
So if it was made by stainless tube and sorbent, here we talk about how to choose the right column will base on what kind of stainless tube and packing material we gotta choose?
How to choose the right packing material?
1. Choose the right packing material to separate will be based on what the properties of the chemical are and what type of chromatography will be.
First let’s talk about the size of diameter of stainless column, it has inner diameter and outer diameter to consider about
The function of different inner diameter column as below
2.1mm and 3.0mm inner size most use for highest performance liquid analyst called UHPLC ( short time and high performance )
4.6mm inner size most use for normal high performance liquid analyst Called HPLC. For 10.0-300mm inner size usually for preparation chromatography called Prep HPLC
Second, let’s talk about the length of column (stainless tube)
Advantages of different column length columns: UHPLC general column length is 50-100mm; in conventional HPLC and prep HPLC, 50-100mm column length is suitable for easy separation of samples, and the analysis time is short. The 150-300mm column length is suitable for samples that are difficult to separate or require high resolution and have a long analysis time. The longer the column length, the higher the column efficiency, but due to the limitation of the packing technology, the column efficiency will be relatively lower than the length of 300 mm or more, so there is rarely a column length greater than 300 mm.
Microscopically, the specifications of column packings material include particle size, pore size, specific surface area, and degree of sealing.
A. The particle size is generally 1.7 μm, 2.7 μm, 3.5 μm, 5 μm, 10 μm, 15 μm, 20 μm, 20-40 μm, 40-60 μm. Among them, 1.7-3.5μm is mainly used for UHPLC rapid analysis, 5μm packing is mainly used for conventional HPLC, 10μm-20μm is usually used for high pressure preparative chromatography (prep HPLC), 20-40μm and 40-60μm are uneven particle size fillers, used for medium Low pressure preparative chromatography (MPLC).
particle size smaller, the column pressure higher and the resolution and sensitivity will be higher
B. Aperture: The size of the surface of the filler particles and the pores inside. 100Å, 150Å, 200Å, 300Å (more suitable for separating macromolecular compounds with large spatial structures such as nucleotides, proteins, etc.).
Specific surface area: The sum of the surface area of the filler particles per unit and the internal pores more bigger. The larger the specific surface area will be bigger, the more contact area with the sample, sample loading will be bigger, the stronger the retention of the sample, and the higher the column efficiency.
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