Background: Danggui Buxue Tang (DBT), a Chinese herbal decoction containing Astragali Radix (AR; roots of Astragalus memebranaceus (Fisch.) Bunge var. mongholicus (Bunge) Hsiao) and Angelicae Sinensis Radix (ASR; roots of Angelica sinensis Oliv.) at a weight ratio of 5:1, is used to improve menopausal syndromes in women. Several lines of evidence indicate that DBT has strong estrogenic property; however, the action mechanism of this herbal decoction is not known. Calycosin, a major flavonoid in AR, shares similar structure with beta-estradiol, and thus which is hypothesized to be the key compound of DBT in responsible for such estrogenic properties. Aims: We aimed to determine the role of calycosin in DBT in terms of its estrogenic functions by the creation of calycosin-depleted DBT (DBT Delta cal) and calycosin-added DBT (DBT+cal) herbal extracts. Methods: The signalings triggered by DBT Delta cal, DBT+cal, and parental DBT were compared in cultured MCF-7 cells by determining: (i) the activation of estrogen responsive element; (ii) the phosphorylation of estrogen receptor a (ER alpha); and (iii) the phosphorylation of Erk1/2. The DBT-induced responses were in dose-and/or time-dependent manners. Results: The estrogenic signals triggered by DBT were markedly reduced in DBT Delta cal, and in contrast the addition of calycosin in DBT, i.e. DBT+cal, enhanced the responses by 2-5 folds; however, calycosin alone did not show such properties. In parallel, the DBT-induced responses could be significantly blocked by inhibitors for estrogen receptor and mitogen activated protein kinases. Conclusion: Thus, we hypothesize that calycosin is an indispensable chemical in DBT, and which plays a linker in orchestrating multi-components of DBT as to achieve the maximal estrogenic functions. These discoveries should be invaluable in drug development and in investigating the modernization of traditional Chinese medicine from a new perspective. (C) 2016 Elsevier Ireland Ltd. All rights reserved.