To study the synergistic effect of polysaccharide from vinegar-baked radix bupleuri(VBCP) with lamivudine(LAM) against hepatitis B virus(HBV) and investigate the mechanism of action.Methods:VBCP，LAM，and VBCP combined with LAM of different concentrations were added into HepG2.2.15 cells.At the same time，the control group was set up and incubated for 48 h.The secretion levels of hepatitis B surface antigen(HBsAg) and hepatitis B e antigen(HBeAg) in the supernatant of the cells were detected by enzyme-linked immunosorbent assay kit，and fluorescence probe quantitative polymerase chain reaction(PCR) was used to detect HBV-deoxyribonucleic acid(DNA) expression in cells.Quantitative analysis of the synergistic effect of polysaccharide from VBCP was conducted by Jin(Zhengjun) formula.The intracellular LAM content was determined by high-performance liquid chromatography.The expression levels of organic cation transporter(OCT) 1，OCT2，P-glycoprotein(P-gp)，and multidrug resistance protein 2(MRP2) were determined by Western blot.Results:Compared with mono-LAM treatment，polysaccharide from VBCP increased the inhibitory effect of LAM on HBsAg secretion，showing an additive effect，and the effect on HBeAg showed a synergistic enhancement.Q value was 6.55，and the inhibitory effect on HBV-DNA was additive.Low-dose，low and middle-dose，and low and high-dose groups of polysaccharide from VBCP could significantly promote the uptake of LAM.The expression of P-gp was significantly decreased in the combined group with a high dose of polysaccharide from VBCP，and groups with polysaccharide from VBCP alone and VBCP combined with LAM significantly increased the expression of OCT1.Conclusion:Polysaccharide from VBCP can play a synergistic anti-HBV effect by increasing the uptake of LAM，and its mechanism may be related to P-gp and OCT1.