Procurement and Selection of Black Cohosh and E. purpurea Samples for Chemical and Biological Analysis

Multiple black cohosh and E. purpurea (Table 1 and Supplementary Tables 1 and 2) samples and reference materials were procured. These included various unfinished products (ie, bulk material that serves as the source for finished products), a limited number of finished products (commercially available tablets or capsules), and reference materials. Currently, NIST SRM are not available for black cohosh or E. purpurea. Therefore, extract reference materials (XRM) and vouchered botanical reference materials (VBRM) were purchased from ChromaDex (Irvine, California; Supplementary Tables 1 and 2). Although all samples were assessed in nontargeted chemical analysis, only a subset of samples were evaluated in the in vitro human hepatocyte nuclear receptor assay. The intent of the in vitro assay was to provide a measure of biological activity for test article selection purposes, therefore only unfinished samples were included.

Summary of Botanical Products Used for Sufficient Similarity Assessment

Abbreviations: XRM, extract reference material; VBRM, verified botanical reference material.

A total of 17 unfinished black cohosh samples, including the NTP test article, were purchased from 8 suppliers (BC1, BC A-P; Table 1 and Supplementary Table 1). Ten finished products (BC Q-Z; Table 1 and Supplementary Table 1) were obtained from 10 manufacturers and contained varied amounts (20–600 mg) of black cohosh according to their labels. Reference material for black cohosh and other cohoshes commonly found as adulterants in black cohosh were purchased from ChromaDex for comparison in this study: black cohosh root XRM (BC AA), Chinese cohosh root VBRM (Cimicifuga dahurica; BC AB), red cohosh root VBRM (Actaea rubra; BC AC), and yellow cohosh root VBRM (Actaea podocarpa; BC AD). All the samples were stored at −20°C.

A similar approach was taken for the procurement of E. purpurea unfinished samples, finished products, and reference materials (details provided in Supplementary Table 2). A total of 13 unfinished samples of E. purpurea, including the NTP test article, were purchased from 8 suppliers (EP1, EP A-L; Table 1 and Supplementary Table 2). Finished products (EP M-Q; Table 1 and Supplementary Table 2) were obtained from 5 suppliers and contained varied amounts (400–1000 mg) of E. purpurea according to their labels. A variety of reference materials were purchased from ChromaDex to better understand whether any observed differences could be related to the plant part or adulteration with other Echinacea species: E. purpurea root XRM (EP R), E. purpurea root VBRM (EP S), E. purpurea leaf and stem VBRM (EP T), and E. purpurea flower VBRM (EP U). Reference material from 2 other Echinacea species were also included: Echinacea angustifolia root XRM (EP V) and Echinacea pallida root VBRM (EP W). All the samples were stored at −20°C.