Suppressed effects of Phyllanthus urinaria L. ethyl acetate extract on hepatitis B virus both in vitro and in vivo

Background: Phyllanthus urinaria L. ( P. urinaria ) extract (PUE) has been used to inhibit hepatitis B virus (HBV). However, the underlying mechanism remains unclear. To investigate which PUE fractions and main components lead to against HBV and approach the relevant molecular mechanisms. Methods: P. urinaria was extracted with water, and then the decoction was extracted by petroleum ether, ethyl acetate, and n-butanol in turn. The HepG2.2.15 cell was treated with aqueous fraction, petroleum ether fraction, ethyl acetate fraction and n-butanol fraction, gallic acid (GA, C 7 H 6 O 5 ) and corilagin (CL, C 27 H 22 O 18 ), respectively. The medium was collected for hepatitis B surface antigen (HBsAg) and hepatitis B e antigen assays. Cell counting kit-8 method was used to identify cell proliferation. Also, the levels of cellular oxygen consumption, reactive oxygen species, and reduced glutathione were detected. The HBV modeling mice were treated with ethyl acetate fraction, entecavir and physiological saline, respectively. The serum was collected for HBsAg and inflammatory cytokines assays. Liver tissue metabolites were screened by LC-MS/MS method. Results: The ethyl acetate fraction (EAF) of P. urinaria could significantly inhibit HBV secretion in HepG2.2.15 ( P < 0.05). Furthermore, two main constitutes in ethyl acetate fraction, GA and CL, could significantly inhibit HBV secretion and reduced cell proliferation ( P < 0.05). Also, GA and CL could increase cellular oxygen consumption, intracellular superoxide anions level, superoxide dismutase level and glutathione depletion. Compared with the Modeling group, EAF significantly decreased the expression levels of HBsAg, IL-1β, IFN-α ( P < 0.05). LC-MS/MS analysis results showed that EAF dramatically up-regulate hydroxyproline, maltotriose, betaine and down-regulate glutathione disulfide, taurocholate, taurochenodeoxycholate ( P < 0.05). Kyoto Encyclopedia of Genes and Genomes results show that the differential metabolites were mainly enriched in ATP-binding cassette transporters pathway. Conclusions: P. urinaria exhibits suppressed effects on HBV by modulating reactive oxygen species formation or metabolomics both in vitro and in vivo. These data indicate that P. urinaria may be an alternative therapeutic agent for the treatment of HBV-related hepatitis.