Strategies for quantitative analysis and preparation of hepatoprotective rosmarinic acid of Lycopus lucidus aerial parts.

The present study investigated strategies for quantifying and preparing rosmarinic acid from Lycopus lucidus, a promising natural source of the bioactive compound. Quantitative 1H nuclear magnetic resonance (qHNMR) was introduced for quantitative analysis, with universal detection being one of its strong features, which made it advantageous for analyzing crude extracts containing various types of compounds. As a result, the L. lucidus leaf extract (LLE) was determined to contain 3.92 ± 0.02% (w/w) rosmarinic acid in the use of caffeine as a universal internal calibrant. Moreover, the countercurrent separation technique was employed to prepare rosmarinic acid from the LLE, especially to effectively remove chlorophylls. Rosmarinic acid was successfully obtained through two rounds of centrifugal partition chromatography, with a purity of 97.92%, as determined by qHNMR. The biological assessment of the LLE, its solvent partitions, and rosmarinic acid revealed that the ethyl acetate-soluble fraction and rosmarinic acid exhibited greater ROS scavenging effects in HepG2 cells than luteolin, a positive control, with the hepatoprotective effect being relevant to the ROS scavenging effect of the major active component, rosmarinic acid.