How to choose the correct column?

Date:2021-11-04 17:32:46 Thursday

Summary:　① Chromatographic column packing matrix　　A. Silica gel matrix: high purity, low cost, high strength, easy chemical modification, but the pH range is limited. Most silica matrix fillers are stable between pH 2-8, but the specially modified silica gel......

　① Chromatographic column packing matrix
　　A. Silica gel matrix: high purity, low cost, high strength, easy chemical modification, but the pH range is limited. Most silica matrix fillers are stable between pH 2-8, but the specially modified silica gel bonded phase can be stabilized at pH 1.5-10.
　　B. Polymer matrix: wide application pH range, stable temperature (high temperature can reach above 80 degrees), and low mechanical strength.
　　②Particle shape
　　 Most modern HPLC fillers are spherical particles, but sometimes irregular particles. Spherical particles provide lower column pressure, higher column efficiency and stability, and longer column life. When using a high-viscosity active phase; irregular particles have a larger specific surface area and a relatively low price.
　　③Particle size
　　The smaller the particle size, the higher the efficiency and the higher the resolution, but at the same time it will lead to a higher column pressure drop. Choose 1.5-3μm packing material to solve some complex samples, UPLC can use 1.5μm packing; another 10μm or larger particle size packing is used as semi-preparative or preparative column.
　　④Carbon content
　　 The carbon content of a chromatographic column refers to the ratio of the bonded phase on the surface of the silica gel, which is related to the specific surface area and bonding coverage. High carbon content improves column capacity, resolution and analysis time, and is used for complex samples that require high resolution; low carbon content has a short analysis time and exhibits different selectivity, and is used for rapid analysis of simple samples and high water-containing active phases. Condition samples. Generally, the carbon content of C18 ranges from 7-19%.
　　⑤ Pore size and specific surface area
　　 HPLC adsorption medium is porous particles, and most of the reaction surface is in the pores. Therefore, molecules must enter the pores to be adsorbed and separated.
　　 Chromatographic column pore size and specific surface area are two complementary concepts. The pore size is small and the specific surface area is large, and vice versa. Large specific surface area, increase the reaction between the sample and the bonded phase, increase storage, sample loading, and separation of complex components; small specific surface area, fast equilibration time, suitable for gradient analysis.
　　⑥ Pore volume and mechanical strength
　　 pore volume, also known as "pore volume". Refers to the size of the void volume per particle. It can well reflect the mechanical strength of the filler. Fillers with large pore volume have slightly weaker mechanical strength than those with small pore volume. Fillers with a pore volume of 1.5 mL/g or less are mostly used for HPLC separation, while fillers with a pore volume greater than 1.5 mL/g are used for size exclusion chromatography and low pressure chromatography.
　　⑦ end cap
　　 Column end capping can reduce the tailing peaks of polar basic compounds due to the interaction with exposed silanol groups. The unblocked bonded phase will have different selectivity compared to the blocked bonded phase, especially for polar samples.