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HPLC determination of ursolic acid in Patrinia scabiosifolia Fisch. ex Trevir. and its inhibitory effect on hela cells

To establish a method for determining ursolic acid content in Patrinia scabiosifolia Fisch. ex Trevir.; and to investigate the inhibitory effect of Patrinia scabiosifolia Fisch. ex Trevir. extract on in vitro cultured human cervical carcinoma Hela cells. HPLC with Agilent XBP-C18 column (4.6 mm × 250 mm, 5 μm) was used. Mobile phase was acetonitrile-methanol-0.5% ammonium acetate solution (68:10:22), while detection wavelength was 210 nm. Human cervical carcinoma Hela cells were cultured in vitro and treated with 0.125-2.0 mg/ml of Patrinia scabiosifolia Fisch. ex Trevir. extracts for 24-72 h. Then, cell proliferation was assayed by MTT assay, apoptosis was detected by FCM, and morphology was observed by TEM. Methodological results showed that regression equation was Y=4.7659x+17.933, R2=0.9998. Ursolic acid presented a good linearity within a 5.21-130.25 μg/ml concentration range. Average recovery was 99.9%, RSD=2.06%. Patrinia scabiosifolia Fisch. ex Trevir. inhibited Hela cells in a dose- and time-dependent manner, with 24 h IC50 being 1.231 mg/ml. FCM revealed increased apoptosis of cells after treatment with Patrinia scabiosifolia Fisch. ex Trevir., while TEM observation found early apoptotic changes in partial cells. With advantages such as simplicity, fastness and good reproducibility, the method proposed can be used for quantitative determination of ursolic acid in Patrinia scabiosifolia Fisch. ex Trevir. Patrinia scabiosifolia Fisch. ex Trevir. has a pronounced inhibitory effect on Hela cells, and induction of apoptosis may be one of its mechanisms.

Author(s): Wei Chen, Yan Liu, Fang Zhao, Lanfang Wu, Yunxi Cai