Chiral chromatographic columns mainly include liquid chiral chromatographic columns and gas chiral chromatographic columns. Liquid chiral chromatographic columns can be divided into natural products, including natural product modifications, and polymers. Natural product-derived chiral chromatographic columns mainly include polysaccharide derivatives such as amylose and cellulose, oligosaccharides such as cyclodextrins and their derivatives, glycoproteins, macrolide antibiotics, and crown ethers. In addition, there are some special purpose ion chiral chromatographic columns. Among them, the chiral column of polysaccharide and its derivatives is a general-purpose chiral column, and it is also the most widely used chiral column at present.
Fig. 1. Structural differences between amylose and cellulose derivatives.
Chiral columns of polysaccharide derivatives are the earliest analytical chromatography for chiral compounds. As a universal chromatographic column for chiral separation, it is estimated that 80-95 % of compounds can be successfully separated by using the polysaccharide derivative chiral column matching method development strategy. Chiral columns can be further classified into coated chiral columns and bonded chiral columns.
Coated polysaccharide derivative chiral column belongs to the first generation of polysaccharide chiral column, and its basic composition includes column silica gel matrix and polysaccharide derivative. The difference in the coated polysaccharide derivative chiral column comes from the difference in the type of polysaccharide and the selective ligand of the derivative (Fig. 1 and Fig. 2).
Fig. 2. Chiral selective ligands for polysaccharide columns.
The commonly used coated chiral chromatographic columns are AD-H, AS-H, OD-H, OJ-H, and the ligand structure is shown in Fig. 3. The selective ligands of OD-H and AD-H are both 3,5-dimethylphenylcarbamate, and the difference in the selectivity of the two to chiral compounds mainly comes from the interaction between matrix and polysaccharide.
Fig. 3. Chiral selective ligands used in daicel coated columns.
The separation principle of the coated polysaccharide derivative chiral chromatography column is to dissolve the polysaccharide derivative, such as THF solvent, and then uniformly coat the dissolved polysaccharide derivative on the porous silica gel matrix. Therefore, the chiral column for coated polysaccharide derivatives has strict requirements on the type of mobile phase, that is, acetone, chloroform, dichloromethane, ethyl acetate and other solvents that can elute the chiral coating layer cannot be used, because even small amounts of these solvents can cause irreversible damage to such chiral column.
The solvents used in coated polysaccharide derivatives chiral column are mainly alkanes, with a certain proportion of alcohol solvents, such as ethanol, isopropanol, n-butanol, etc. When analyzing polar compounds such as acid and alkali, a certain proportion of organic acid and alkali additives can also be added. In addition, polar elution mode can also be used, that is, 100% organic alcohols or acetonitrile and its mixture are used as the mobile phase, but reversed-phase elution mode cannot be used. The reversed-phase elution mode can use another type of coated chiral column, including AD-RH, OD-RH, OJ-RH, AS-RH, etc.
As the second-generation polysaccharide derivative chiral column, the polysaccharide derivative of the bonded chiral column is chemically bonded to the silica gel matrix of the chromatographic column stationary phas, thereby improving the tolerance to non-standard mobile phases. Bonded chiral columns expand the use of polysaccharide derivative chiral columns to the extent that there is no solvent limitation. Bonded chiral columns not only give full play to the column selectivity, but also give full play to the chiral selectivity of the mobile phase.
Fig. 4. Chiral ligands for immobilized columns of daicel.
Common bonded chiral chromatographic columns include IA, IB, IC, ID, IE, IF and the latest IG, and the corresponding ligand structures are shown in Fig. 4. Among them, IB and IC are cellulose-type polysaccharide derivatives, and the rest are amylose-type polysaccharide derivatives. Generally, there is overlap in the selective ligands for bonded chiral columns and coated chiral columns, but there are also differences. For example, chiral ligands such as 3,5-dichlorophenylcarbamate and 3-chloro-4-methylphenylcarbamate are not found in coated chiral columns. Compared with coated polysaccharide derivative chiral columns, bonded polysaccharide chiral columns can use various non-standard solvents as mobile phases, such as dichloromethane, tetrahydrofuran, ethyl acetate, which is the mobile phase prohibited by the coated chromatographic column. In addition, when chiral compounds are resolved by a polysaccharide chiral column, the π-π interaction, hydrogen bond interaction, dipole interaction and inclusion in the system will affect the separation effect.
Whether it is gas chromatography or liquid chromatography, chiral chromatographic columns of cyclodextrin and its derivatives mainly use β- or γ-cyclodextrins. Generally, the ligands of gas-phase cyclodextrin chiral chromatography columns and liquid-phase cyclodextrin chiral chromatography columns are different. Fig. 5 shows several common cyclodextrin ligands, of which the first three gas phase are used more, and the latter three liquid phases are used more.
Fig. 5. Common ligands for cyclodextrin derivative chiral columns.
In the resolution of chiral compounds, the main interactions of cyclodextrin chiral columns include surface hydrogen bonding interactions and inclusion and electrostatic interactions in the cyclodextrin cavity. Common liquid-phase cyclodextrin chiral columns mainly include Cyclobond series: Cyclobond I 2000, Cyclobond I 2000 AC, Cyclobond I 2000 DM, Cyclobond II, Cyclobond II AC, etc.
Different chiral stationary phases may have different chiral interaction sites and types of forces, including ionization interactions, hydrogen bond interactions, π-π interactions, including mutual and electrostatic interactions, etc. Pirkle-type chiral columns are representative of π-π interactions, and these chiral columns are more suitable for separating chiral compounds containing benzene rings or other heterocyclic types.
Protein chiral chromatographic columns mainly include HSA, TAG, and CBH. Among them, TAG has a wide range of applications, HSA is suitable for strongly acidic compounds, and CBH is suitable for strongly basic compounds. The resolution mechanism of chiral compounds by protein chiral chromatography columns is more similar to the "lock-key" model except for hydrogen bond interactions and ionization interactions. Typically, protein chiral columns are only used in reversed-phase mode.
When using liquid chromatography to separate chiral compounds, the choice of chiral chromatographic column is more important than the mobile phase, which is more important than the choice of operating conditions. BOC Sciences has experienced chiral chromatography experts to provide cost-effective screening services. If you are interested in our chiral column screening services, please contact us for more information.
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