Abstract:

Organic impurity determination is a critical aspect since it has a great impact on the assigned content of chemical reference standards (CRS). This is especially the case for CRS of natural products (CRSNPs) which are mostly obtained from medicinal plants or animals. Total amount of organic impurities of CRSNPs is generally determined by high performance liquid chromatography based on official or reported methods. Since the brand of column is not always prescribed or when the brand prescribed is not available in the laboratory, this can cause serious consequences, especially when the sample contains a basic or weak acid compound with pK_a values ranged from 3 to 6. In this paper, columns were originally selected based on a peer-reviewed column characterization database. Firstly two columns with orthogonal selectivity (selectivity factor F≥6) were selected from the database to determine the purity of the respective CRSNPs. The selection was performed to minimize the risk to obtain different values caused by use of various brands of columns. If there is no significant difference between their results, then the purity result is cross-validated. Otherwise, a third column has to be selected from the database and its selectivity should be similar to the above one which give better separation (F≤2). The approach has been illustrated by the purity determinations of N-trans-p-coumaroyltyramine and (-)-epicatechin-3-O-gallate CRSNPs, and it is most likely to be applied in more CRSNPs, especially those of basic or weak acid ones.

Key words: (-)-epicatechin-3-O-gallate, N-trans-p-coumaroyltyramine, chemical reference standards of natural products (CRSNPs), chromatographic purity, column characterization database, rational column selection approach