Direct determination of astragalosides and isoflavonoids from fresh Astragalus membranaceus hairy root cultures by high speed homogenization coupled with cavitation-accelerated extraction followed by liquid chromatography-tandem mass spectrometry

Abstract

A direct analysis approach for plant in vitro cultures, namely, high speed homogenization coupled with cavitation-accelerated extraction (HSH-CAE) followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), was developed for the simultaneous determination of six astragalosides and five isoflavonoids in Astragalus membranaceus hairy root cultures (AMHRCs). In comparison to reported Soxhlet extraction (SE) and ultrasound-assisted extraction (UAE) methods, the proposed sample preparation procedure (HSH-CAE) offers significant improvements with regard to simplicity in operation (elimination of biomass drying and grinding), high efficiency, enhanced yield and green aspects in terms of saving energy cost and minimizing the generation of waste. In addition, the HSH-CAE mechanism was clarified via cytohistological studies of samples at cellular/tissular levels. Moreover, the established LC-MS/MS method provided linearity with correlation coefficients above 0.9991, limit of detections (LODs) below 1.77 ng mL⁻¹, relative standard deviations (RSDs) below 6.01%, and recoveries above 96.84%. Furthermore, the proposed HSH-CAE-LC-MS/MS method was also successfully applied for screening high-productive AMHRCs. Overall, this study opened up a new avenue for the direct determination of secondary metabolic profiles from fresh plant in vitro cultures, which was valuable for improving the quality control of plant cell/organ cultures and shed light on the metabolomics analysis from biological samples.