How to avoid the tailing problem of basic compounds in HPLC analysis?

For the Pharmaceutical workers, Alkaloids it is no a unfamiliar thing that is one of the most important and valuable biologically active ingredients in Chinese herbal medicine. Alkaloids can be divided to 59 species. They have complex structure that cannot separation well by tradition method. Before the HPLC, the separation of alkaloids it is a problem for scholars. The most obvious problem in the analysis of basic compounds is peak tailing, and the stronger the alkalinity, the more likely it is that tailing will occur.

Let’s learn more about how to avoid the peek tailing in HPLC!

1. Mobile phase

The phenomenon of peak trailing can be improved by adjusting the mobile phase. In the early stage, the pH value range of the mobile phase should be considered when selecting the mobile phase. For basic compounds, the pH of the mobile phase will have a greater impact on the peak shape, as well as the retention factor and selectivity. UHPLCS suggests select PH≤2.5 if you want the separation of basic compounds that can be formed in a good peak shape. The PH steady value of the UHPLCS Liquid chromatography column is 1.5-11. It is suitable for liquid phase analysis of various needs and various separation modes.

2. stationary phase

C18 chromatographic column is a column often used in reversed-phase chromatographic analysis. However, the octadecyl carbon chain is very long, and the steric hindrance is very large when bonding, resulting in a low bonding rate. In this way, a large number of unbonded silyl hydroxyl groups will remain. Silanols are easily dissociated in some reverse mobile phases. Due to the secondary interaction between the free silanol and the dissociated alkaline compound, the peak tailing of the alkaline compound appears. Therefore, when analyzing basic compounds, choosing a bonded phase with low silanol activity is the key to obtaining a good peak shape. For the selection of the stationary phase, it is best to use ultra-pure silica gel with a purity of 99.99% or more. The amount of metal residue is relatively low, which increases the bonding rate and reduces the free silanols.

UHPLCS Liquid chromatography column with High purity silica gel matrix packing. USHB C18-BIO is designed  for the separation of a wide range of alkaline compounds. The pH tolerance range is 1.5-10, metal impurities are extremely low, and the secondary sealing technology is adopted. This is mainly to further reduce the activity of silanol so that a good peak shape can be obtained during analysis.