By modifying the matrix or bonding different groups, different separation types of stationary phases are formed, usually including normal phase, reversed-phase, ion exchange, SEC, chiral, and other stationary phases.
The chromatographic mode in which the polarity of the modified groups on the matrix is greater than the polarity of the mobile phase is called normal phase chromatography, which relies on the different polarity of the sample to distribute between the stationary phase and the mobile phase to achieve separation. In this separation mode, the sample is eluted from the purification column in order from weak to strong polarity.
Commonly used normal phase stationary phases include unmodified silica gel; diol-based, amino, cyano-based, and other silica-based stationary phases; alumina, etc. The normal phase silica gel stationary phase is mainly used in the preliminary purification of synthetic non-polar or medium-polar intermediates. The conventional flow system is n-hexane/ethyl acetate, dichloromethane/methanol, etc.
Its advantages are simple purification and simple sample post-processing; its disadvantages are low separation performance (compared with reversed-phase immobilization), poor reproducibility, and strong adsorption and even irreversible adsorption of certain substances. In addition, some polar groups (diol groups, amino groups, cyano groups, etc.) bonded normal phase stationary phases are often used in reversed phase conditions. For polar compounds that are not strongly retained in reversed phase stationary phases such as C18, they can Obtain a good separation and purification effect.
Alumina stationary phase is divided into three types: acidic, neutral, and basic. Acidic alumina is generally pretreated with acidic solvents. It has weak cationic properties. It is easier to remain neutral and negatively charged substances on the surface, and cannot retain positively charged substances. It is mainly used in acid pigments, aldehydes, and acids.
Compound separation and purification. Alkaline alumina is usually pretreated with an alkaline solution. It has anionic characteristics and cation exchange function. It has a strong adsorption effect on polar cation samples. It is mainly used for the separation and purification of alkaline pigments, alkaloids, and other alkaline substances. Neutral alumina is not sensitive to the acidity and alkalinity of the sample and is mainly used for acid-base unstable compounds such as glycosides, aldehydes, and esters.
The chromatographic mode in which the polarity of the modified groups on the matrix is less than the polarity of the mobile phase is called reversed-phase chromatography. In this separation mode, the order in which the samples are eluted is exactly the opposite of the normal phase mode: To Ruo was eluted in turn. Most of the reversed-phase stationary phases are based on silica gel matrix bonded with different bonded phases such as C18, C8, C4, C1, phenyl, and other stationary phases. They are widely used in the purification and purification of natural products, peptides, and proteins, and have excellent separation effects. , Good reproducibility, and long service life.
Ion exchange chromatography is a separation mode through the interaction between the surface ion charges of the sample molecules and the surface ion charges of the chromatography packing. The surface of the stationary phase is positively charged, and those that retain anions are called anion-exchange fillers; the surface of the stationary phase is negatively charged, and those that retain cations are called cation exchange fillers. In addition, they are classified as strong ions according to the strength of the ion binding state on the filler surface. Exchange packing and weak ion exchange packing.
The common surface groups of cation exchange fillers are sulfonic acid groups (strong cation exchange), phosphoric acid groups, carboxylic acid groups, and phenolic hydroxyl groups; the common surface groups of anion exchange fillers are primary amino groups, secondary amino groups, tertiary amino groups, and quaternary Amine group (strong anion exchange packing). When ion exchange packing is used in preparation and purification, the resin is usually used as the matrix, and there are more silica matrix and non-porous gels in analytical applications due to pressure and resolution considerations.
A chiral stationary phase is a stationary phase in which derivatives such as polysaccharides, cyclodextrins, and proteins are bonded or coated on a substrate such as silica gel, and chiral recognition and resolution of chiral substances are performed to achieve the purpose of separation and purification.
Since chiral stationary phases are very expensive, in order to protect the stationary phase as much as possible, the chemical purity of the sample is generally required. Usually, the sample is initially purified to improve the chemical purity and remove impurities that are likely to damage or adsorb the stationary phase.
Surface Modification/Bonding Phase Of FLASH Purification Column Stationary Phase
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