An integrated method by ultrasonic enhancement in the natural deep eutectic solvents for extraction and separation of chlorogenic acids from Duhaldea nervosa

The application of Natural Deep Eutectic Solvents (NADES) as a sustainable alternative to conventional organic solvents for extraction of bioactive compounds from traditional Chinese medicine (TCM) has gained considerable attention. However, challenges remain in removal of NADES and the recovery of extracted products. This study developed an efficient and eco-friendly ultrasonic-assisted NADES method for the extraction and enrichment of chlorogenic acid components from Duhaldea nervosa (DN). Fifteen NADES types were synthesized and evaluated, among which a mixture of choline chloride and 1,3-butanediol at a molar ratio of 1:6 was identified as optimal. Extraction conditions were optimized using Box-Behnken design combined with Response Surface Methodology (RSM), yielding the following parameters: 64 % NADES content, 11 mL solvent per gram of dry matter, and ultrasonic power of 300 W. The NKA-II resin demonstrated high efficacy in adsorbing and desorbing chlorogenic acids, with 40 % ethanol selected as the optimal eluent. This approach enabled efficient NADES removal and recovery of chlorogenic acids. The ultrasonic-enhanced continuous extraction and enrichment technology within the NADES system offers notable advantages including operational simplicity, environmental friendliness, cost-effectiveness, and high efficiency. A total of 55 chlorogenic acid components in the product obtained from the optimal extraction and enrichment process of DN were identified using feature-based molecular networking analysis. Furthermore, the effects of both the crude ethanol extract of DN and the chlorogenic acid-enriched product on RAW264.7 cell viability and lipopolysaccharide (LPS)-induced nitric oxide (NO) production in macrophages were systematically evaluated.