Background: Root of Panax ginseng C. A. Mey (Renseng in Chinese) is a famous Traditional Chinese Medicine. Ginsenosides are the major bioactive components. However, the shortage and high cost of some ginsenoside reference standards make it is difficult for quality control of P. ginseng. Objective: A method, single standard for determination of multicomponents (SSDMC), was developed for the simultaneous determination of nine ginsenosides in P. ginsen g (ginsenoside Rg1, Re, Rf, Rg2, Rb1, Rc, Rb2, Rb3, Rd). Materials and Methods: The analytes were separated on Inertsil ODS-3 C18 (250 mm × 4.6 mm, 5 μ m) with gradient elution of acetonitrile and water. The flow rate was 1 mL/min and detection wavelength was set at 203 nm. The feasibility and accuracy of SSDMC were checked by the external standard method, and various high-performance liquid chromatographic (HPLC) instruments and chromatographic conditions were investigated to verify its applicability. Using ginsenoside Rg1as the internal reference substance, the contents of other eight ginsenosides were calculated according to conversion factors (F) by HPLC. Results: The method was validated with linearity (r2 ≥ 0.9990), precision (relative standard deviation [RSD] ≤2.9%), accuracy (97.5%–100.8%, RSD ≤ 1.6%), repeatability, and stability. There was no significant difference between the SSDMC method and the external standard method. Conclusion: New SSDMC method could be considered as an ideal mean to analyze the components for which reference standards are not readily available.