Quantitative Analysis and Comparison of Four Major Flavonol Glycosides in the Leaves of Toona sinensis (A. Juss.) Roemer (Chinese Toon) from Various Origins by High‑Performance Liquid Chromatography‑Diode Array Detector and Hierarchical Clustering Analysis

Background: Toona sinensis (A. Juss.) Roemer is an endemic species of Toona genus native to Asian area. Its dried leaves are applied in the treatment of many diseases; however, few investigations have been reported for the quantitative analysis and comparison of major bioactive flavonol glycosides in the leaves harvested from various origins. Objective: To quantitatively analyze four major flavonol glycosides including rutinoside, quercetin‑3‑O‑β‑D‑glucoside, quercetin‑3‑O‑α‑L‑rhamnoside, and kaempferol‑3‑O‑α‑L‑rhamnoside in the leaves from different production sites and classify them according to the content of these glycosides. Materials and Methods: A high‑performance liquid chromatography‑diode array detector (HPLC‑DAD) method for their simultaneous determination was developed and validated for linearity, precision, accuracy, stability, and repeatability. Moreover, the method established was then employed to explore the difference in the content of these four glycosides in raw materials. Finally, a hierarchical clustering analysis was performed to classify 11 voucher specimens. Results: The separation was performed on a Waters XBridge Shield RP18 column (150 mm × 4.6 mm, 3.5 μm) kept at 35°C, and acetonitrile and H2O containing 0.30% trifluoroacetic acid as mobile phase was driven at 1.0 mL/min during the analysis. Ten microliters of solution were injected and 254 nm was selected to monitor the separation. A strong linear relationship between the peak area and concentration of four analytes was observed. And, the method was also validated to be repeatable, stable, precise, and accurate. Conclusion: An efficient and reliable HPLC‑DAD method was established and applied in the assays for the samples from 11 origins successfully. Moreover, the content of those flavonol glycosides varied much among different batches, and the flavonoids could be considered as biomarkers to control the quality of Chinese Toon.