ShenMai, an intravenous injection prepared from steamed Panax ginseng roots (Hongshen) and Ophiopogon japonicus roots (Maidong), is used as an add-on therapy for coronary artery disease and cancer; saponins are its bioactive constituents. Since many saponins inhibit human organic anion-transporting polypeptides (OATP)1B, this investigation determined the inhibition potencies of circulating ShenMai saponins on the transporters and the joint potential of these compounds for ShenMai-drug interaction. Circulating saponins and their pharmacokinetics were characterized in rats receiving a 30-min infusion of ShenMai at 10 mL/kg. Inhibition of human OATP1B1/1B3 and rat Oatp1b2 by the individual saponins was investigated in vitro; the compounds’ joint inhibition was also assessed in vitro and the data was processed using the Chou–Talalay method. Plasma protein binding was assessed by equilibrium dialysis. Altogether, 49 saponins in ShenMai were characterized and graded into: 10–100 μmol/day (compound doses from ShenMai; 7 compounds), 1–10 μmol/day (17 compounds), and <1 μmol/day (25 compounds, including Maidong ophiopogonins). After dosing, circulating saponins were protopanaxadiol-type ginsenosides Rb1, Rb2, Rc, Rd, Ra1, Rg3, Ra2, and Ra3, protopanaxatriol-type ginsenosides Rg1, Re, Rg2, and Rf, and ginsenoside Ro. The protopanaxadiol-type ginsenosides exhibited maximum plasma concentrations of 2.1–46.6 μmol/L, plasma unbound fractions of 0.4–1.0% and terminal half-lives of 15.6–28.5 h (ginsenoside Rg3, 1.9 h), while the other ginsenosides exhibited 0.1–7.7 μmol/L, 20.8–99.2%, and 0.2–0.5 h, respectively. The protopanaxadiol-type ginsenosides, ginsenosides without any sugar attachment at C-20 (except ginsenoside Rf), and ginsenoside Ro inhibited OATP1B3 more potently (IC50, 0.2–3.5 µmol/L) than the other ginsenosides (≥22.6 µmol/L). Inhibition of OATP1B1 by ginsenosides was less potent than OATP1B3 inhibition. Ginsenosides Rb1, Rb2, Rc, Rd, Ro, Ra1, Re, and Rg2 likely contribute the major part of OATP1B3-mediated ShenMai-drug interaction potential, in an additive and time-related manner.