Scale-Up Isolation of High-Purity Shatavarins from <i>Asparagus racemosus</i> via Flash Chromatography with Bioavailability and Pharmacological Insights

Asparagus racemosus, commonly known as Shatavari, is an established Ayurvedic plant whose bioactivity is credited to its steroidal saponins, generally referred to as shatavarins, exerting adaptogenic and estrogenic effects. However, traditional methods of isolation often produce products with limited purity and poor reproducibility, which seriously limit scalability in pharmacological applications. This study describes an effective, scalable, flash chromatography-based approach toward the isolation and purification of six bioactive steroidal saponins, AR-1 to AR-6, from AR root material. The methanolic extract was subjected to successive liquid–liquid partitioning and multistep chromatographic purification to give high-purity compounds identified as spirostan derivatives (AR-1 to AR-4), shatavarins, and aglycone derivatives (AR-5 and AR-6). The structural identities and chemical homogeneity were clearly supported by various spectroscopic analyses: HPLC-ELSD, FTIR, NMR, and LC-MS. The in silico ADME predictions performed by SwissADME were used as a prioritization strategy to inform compound selection for biological testing, providing relative comparisons of absorption potential rather than validating oral bioavailability. Derivatives of aglycone compounds AR-5 and AR-6 showed greater predicted absorption potential, while highly glycosylated saponins AR-2 and AR-4 showed low permeability, as expected for compounds of their size and polarity. While steroidal saponins are not typical of oral drug chemical space, these models correlate well with permeability and metabolic properties. Molecular docking studies of GPER and estrogen receptors (ERα, ERβ, and ERRγ) showed binding affinities of −7.7 to −12.4 kcal/mol, with AR-1, AR-2, and AR-4 showing a preference for hydrophobic and hydrogen-bonding interactions in GPER and ERα binding pockets typical of estrogen ligands. The developed flash chromatography workflow represents an efficient, scalable route to pharmacologically relevant standards of shatavarins. Combined with chromatographic and in silico results, the findings reveal the translational potential of AR saponins as multitarget phytoestrogens for hormone-modulatory and adaptogenic therapeutic applications.