A new validated high-performance liquid chromatography method for standardization of rosmarinic acid in Salvia extracts

Introduction: Tea bags or infuses of Salvia species from Lamiaceae family are traditionally used for the treatment of cough, and throat inflammations. They are known for antioxidant properties mainly related to the presence of rosmarinic acid (RA). Therefore it is necessary to develop a reliable analytical method for RA assay for standardization of Salvia species and also other plants containing RA like Melissa, Origanum, Lavandula, Rosmarinus, Thymus, Zataria, Mentha, Perovskia, Zhumeria, and Satureja species. In this study using a suitable extraction method by removing unwanted components present in crude methanol extract, phenolic content containing RA was extracted from dry powders of six Salvia species. Then, a suitable high-performance liquid chromatographic (HPLC) method was optimized for quantification of RA in Salvia species. Methods: HPLC analysis was done on a Waters system, equipped with 515 HPLC pump and waters 2487 dual wavelength absorbance detector. The column was a Nova-Pak C18 (3.9 × 150 mm), and Millenium software was used for the determination of the compounds and processing the data. The method was validated according to USP 32 requirements. Results: Among the investigated 6 species, S. virgata was the richest in RA level, demonstrating 3.50 ± 0.12 mg/g, followed by S. sclarea and S. chloroleuca showing 1.65 ± 0.08 and 1.65 ± 0.21 mg/g. S. ceratophylla with 0.10 ± 0.01 mg/g of RA in dried plant powder was the poorest. Conclusion: The validated HPLC method allows determination of amounts as low as 2.5 µg/ mL of RA and linearity in the ranges of 2.5-25 µg/mL and 100–600 µg/mL, which is suitable for standardization of Salvia species in traditional and pharmaceutical formulations