Calycosin orchestrates the functions of Danggui Buxue Tang, a Chinese herbal decoction composing of Astragali Radix and Angelica Sinensis Radix: An evaluation by using calycosin-knock out herbal extract

Danggui Buxue Tang (DBT) is a classical Chinese herbal decoction containing two herbs, Astragali Radix (AR) and Angelicae Sinensis Radix (ASR), which serves as dietary supplement for treating women menopausal syndromes. Pharmacological studies indicate that DBT has estrogenic, erythropoietic and osteogenic properties; however, the action mechanism for this complex herbal decoction is not known. Calycosin, a major flavonoid in AR, shares similar structure with β-estradiol, and thus which is hypothesized to be the critical compound of DBT. Here, we aim to investigate the role of calycosin in DBT in terms of its biological functions by using a calycosin-depleted DBT decoction (DBTΔcal). The biological functions of DBTΔcal and parental DBT were systematically compared. In order to standardize DBT decoction, four chemical markers were determined and quantified by HPLC. A semi-preparative HPLC method was utilized to prepare DBTΔcal. The authenticity of DBTΔcal was evaluated by LC-QQQ-MS/MS. To reveal the effect of calycosin on DBT functions, several cell assays related to the known properties of DBT were revealed, including estrogenic, erythropoietic and osteogenic functions. As compared to parental DBT, the estrogenic, erythropoietic and osteogenic abilities were markedly reduced in DBTΔcal. However, calycosin alone did not show significant responses. Our results suggest that calycosin is a bioactive chemical in DBT decoction, and which could play a key linker in orchestrating multi-components of DBT as to achieve maximal functions. These discoveries should be invaluable in drug development and in investigating the modernization of traditional Chinese medicine from a new perspective.