| 摘要 |
[Objectives] To establish an efficient and environmentally friendly separation and purification method for the large-scale preparation of the major active components—eleutherol, eleutherine, and isoeleutherine—from the ethnomedicinal plant Eleutherine americana Merr. et K. Heyne. [Methods] The sample of E. americana bulbs was initially extracted with ethanol, followed by three successive extractions with ethyl acetate-water (2：1, V/V) to obtain the target component-enriched fraction. Eight solvent systems were systematically optimized, and a mixture of petroleum ether-ethyl acetate-ethanol-water (5：5：6：4, V/V/V/V) was identified as the optimal solvent system for high-speed counter-current chromatography (HSCCC) separation under conditions of 900 rpm, 2 mL/min, and 35 ℃. The crude HSCCC product was further purified by silica gel column chromatography (200-300 mesh) using gradient elution with a solvent system of n-hexane-dichloromethane-ethyl acetate (varying from 10：5：1 to 4：5：1, V/V/V). UPLC-PDA (Agilent SB-C18 column) and nuclear magnetic resonance spectroscopy (600 MHz) were comprehensively employed to assess compound purity and confirm molecular structures. [Results] An optimized technique integrating HSCCC and silica gel column chromatography was established, successfully enabling the large-scale preparation of three bioactive components: eleutherol (purity 99%), eleutherine (purity 98%), and isoeleutherine (purity 98%). Structural identification results were consistent with those reported in the literature. Compared to traditional methods, the new approach demonstrated improved separation efficiency and reduced solvent consumption. [Conclusions] The combined separation method utilizing HSCCC and silica gel column chromatography established in this study demonstrates notable advantages, including high efficiency, environmental friendliness, and cost-effectiveness, enabling the large-scale preparation of the three major active components from E. americana. This approach outperforms conventional methods by offering higher separation efficiency, reduced solvent consumption, and superior product purity, providing a robust technical solution for the development and utilization of bioactive compounds from E. americana. Moreover, it offers a novel methodological reference for the isolation and purification of other natural products. |
