Sepaflash ® C18 reversed phase purification column application case -- Separation and purification of CIS and trans isomers

In the process of chemical synthesis of pharmaceutical intermediates, many isomers, including CIS and trans isomers, are synthesized at the same time by applied technology research center CIS trans isomers belong to one of the stereoisomers The reason is that the two carbon atoms connected by double bonds can not rotate freely around the σ bond The chemical properties of a pair of CIS and trans isomers are basically the same because of the same connection order of atoms or atomic groups and the same position of double bonds, but with different spatial arrangement However, the physical properties of CIS and trans isomers are different, such as melting point, solubility, etc., so some CIS and trans isomers can be separated by vacuum distillation and recrystallization, while some CIS and trans isomers can be separated by column chromatography In this paper, a pair of CIS and trans isomers were obtained in the process of synthesis The purification method of vacuum distillation and recrystallization could not meet the purity requirements Based on this situation, the purification personnel tried to use the rapid preparation of liquid chromatography sepabean ® machine with sepaflash ® series reversed-phase purification column to separate and purify the samples Experimental part 1 Sample information the structural formula of the sample in this experiment is shown in Figure 1, in which the target of the separation and purification experiment is cis isomer and the impurity is trans isomer Figure 1 Structural formula of the sample: A is trans isomer, B is cis isomer 2 Preliminary analysis of the sample Preliminary purity analysis of the sample was carried out by HPLC (see Figure 2) The result shows that the purity of the sample is 79%, and the main impurity is trans isomer in the sample According to the analysis of HPLC, the target product and impurities were separated by baseline, so the subsequent separation and purification were carried out by reverse phase separation method Fig 2 HPLC analysis spectrum of the sample Fig 3 Flash preparation and purification experimental parameters of the sample are shown in Table 1, and flash preparation and purification spectrum is shown in Fig 3 Table 1 Sepabean ® machine separation column 330 g sepaflash ® bonded series C18 reversed phase purification column (spherical silica gel, 20-45 μ m, 100 μ m, Order No.: sw-52 22-330-sp) detection wavelength 220 nm; 254 nm mobile phase solvent A: water; solvent B: methanol flow rate 40 ml / min injection volume 5 ml (1.4g) elution gradient time (min) solvent B (%) 0 60 35 90 80 90 Figure 3 Flash preparation and purification spectrum of sample 4 Purity analysis of purified product After the prepared sample is evaporated and dried, the purity of the sample is analyzed by HPLC (see Figure 4) The result shows that the purity is 98.7%, which meets the purity standard of the target product It should be noted that in the HPLC analysis and detection of the purified product, the separation time is shortened by optimizing the method Therefore, compared with Fig 2 and Fig 4, it can be found that the retention time of impurities and target products in Fig 4 are shortened Figure 4 Results and discussion of HPLC analysis spectrum of purified products For the separation and purification of isomers, researchers usually use chiral separation column or high-pressure preparation column with high-pressure preparation liquid chromatography for preparation and purification, but the cost of such methods is very painful The application cases in this paper show that for some isomer samples which are not difficult to separate, medium pressure preparation and separation can also achieve a good separation and purification effect, which will undoubtedly greatly reduce the preparation cost of researchers and obtain better benefits Sepabean ® machine produced by Santai technology, in combination with sepaflash ® series C18 reversed-phase purification column, can provide users with fast and efficient sample preparation and purification solutions, and help laboratory researchers easily complete various sample purification tasks To learn more about the detailed specifications of sepabean ® machine, or to order the flash purification column used with it, please visit CBG online store.